Co-reporter:Brian F. Fisher, Seong Ho Hong, and Samuel H. Gellman
Journal of the American Chemical Society September 27, 2017 Volume 139(Issue 38) pp:13292-13292
Publication Date(Web):September 12, 2017
DOI:10.1021/jacs.7b07930
We describe the use of thioester exchange equilibria to measure the propensities of amino acid residues to participate in helical secondary structure at room temperature in the absence of denaturants. Thermally or chemically induced unfolding has previously been employed to measure α-helix propensities among proteinogenic α-amino acid residues, and quantitative comparison with precedents indicates that the thioester exchange system is reliable for residues that lack side chain charge. This system allows the measurement of α-helix propensities for d-α-amino acid residues and propensities of residues with nonproteinogenic backbones, such as those derived from a β-amino acid, to participate in an α-helix-like secondary structure.
Co-reporter:Nicole C. Thomas, Gail J. Bartlett, Derek N. Woolfson, and Samuel H. Gellman
Journal of the American Chemical Society November 22, 2017 Volume 139(Issue 46) pp:16434-16434
Publication Date(Web):November 8, 2017
DOI:10.1021/jacs.7b07225
The formation and deposition of amyloids is associated with many diseases. β-Sheet secondary structure is a common feature of amyloids, but the packing of sheets against one another is distinctive relative to soluble proteins. Standard methods that rely on perturbing a polypeptide’s sequence and evaluating impact on folding can be problematic for amyloid aggregates because a single sequence can adopt multiple conformations and diverse packing arrangements. We describe initial steps toward a minimum-sized, soluble model system for the amyloid state that supports comparisons among sequence variants. Critical to this goal is development of a new linking strategy to enable intersheet association mediated by side chain interactions, which is characteristic of the amyloid state. The linker design we identified should ultimately support exploration of relationships between sequence and amyloid state stability for specific strand-association modes.
Co-reporter:Melissa J. MacDonald, Natasha R. Cornejo, and Samuel H. Gellman
ACS Macro Letters July 18, 2017 Volume 6(Issue 7) pp:695-695
Publication Date(Web):June 19, 2017
DOI:10.1021/acsmacrolett.7b00396
Nontoxic cryoprotectants are needed for storage of tissues and food preservation. Frozen tissue is particularly susceptible to damage caused by formation of large ice crystals during the thawing process. The current practice of using 5 wt % DMSO for cryopreservation does not produce 100% cell viability post-thaw, at least in part because of DMSO toxicity that is manifested during the freezing and thawing stages of the process. Recently, poly(vinyl alcohol) (PVA) has shown promise in inhibiting ice recrystallization, an activity that is critical for cryoprotection. Inspired by this discovery, we have evaluated nylon-3 polymers for ice recrystallization inhibition activity and for toxicity toward mammalian cells. A survey of homo- and heteropolymers, with side chains bearing variable functionality, has identified new nylon-3 materials that display excellent ice recrystallization inhibition activity and low toxicity.
Co-reporter:Dr. James W. Checco; Samuel H. Gellman
ChemBioChem 2017 Volume 18(Issue 3) pp:291-299
Publication Date(Web):2017/02/01
DOI:10.1002/cbic.201600545
AbstractInhibition of specific protein–protein interactions is attractive for a range of therapeutic applications, but the large and irregularly shaped contact surfaces involved in many such interactions make it challenging to design synthetic antagonists. Here, we describe the development of backbone-modified peptides containing both α- and β-amino acid residues (α/β-peptides) that target the receptor-binding surface of vascular endothelial growth factor (VEGF). Our approach is based on the Z-domain, which adopts a three-helix bundle tertiary structure. We show how a two-helix “mini-Z-domain” can be modified to contain β and other nonproteinogenic residues while retaining the target-binding epitope by using iterative unnatural residue incorporation. The resulting α/β-peptides are less susceptible to proteolysis than is their parent α-peptide, and some of these α/β-peptides match the full-length Z-domain in terms of affinity for receptor-recognition surfaces on the VEGF homodimer.
Co-reporter:Brian F. Fisher and Samuel H. Gellman
Journal of the American Chemical Society 2016 Volume 138(Issue 34) pp:10766-10769
Publication Date(Web):August 16, 2016
DOI:10.1021/jacs.6b06177
α/γ-Peptide foldamers containing either γ4-amino acid residues or ring-constrained γ-amino acid residues have been reported to adopt 12-helical secondary structure in nonpolar solvents and in the solid state. These observations have engendered speculation that the seemingly flexible γ4 residues have a high intrinsic helical propensity and that residue-based preorganization may not significantly stabilize the 12-helical conformation. However, the prior studies were conducted in environments that favor intramolecular H-bond formation. Here, we use 2D-NMR to compare the ability of γ4 residues and cyclic γ residues to support 12-helix formation in more challenging environments, methanol and water. Both γ residue types support 12-helical folding in methanol, but only the cyclically constrained γ residues promote helicity in water. These results demonstrate the importance of residue-based preorganization strategies for achieving stable folding among short foldamers in aqueous solution.
Co-reporter:Marlies V. Hager, Lisa M. Johnson, Denise Wootten, Patrick M. Sexton, and Samuel H. Gellman
Journal of the American Chemical Society 2016 Volume 138(Issue 45) pp:14970-14979
Publication Date(Web):November 4, 2016
DOI:10.1021/jacs.6b08323
Activation of a G protein-coupled receptor (GPCR) causes recruitment of multiple intracellular proteins, each of which can activate distinct signaling pathways. This complexity has engendered interest in agonists that preferentially stimulate subsets among the natural signaling pathways (“biased agonists”). We have examined analogues of glucagon-like peptide-1 (GLP-1) containing β-amino acid residues in place of native α residues at selected sites and found that some analogues differ from GLP-1 in terms of their relative abilities to promote G protein activation (as monitored via cAMP production) versus β-arrestin recruitment (as monitored via BRET assays). The α → β replacements generally cause modest declines in stimulation of cAMP production and β-arrestin recruitment, but for some replacement sets cAMP production is more strongly affected than is β-arrestin recruitment. The central portion of GLP-1 appears to be critical for achieving bias toward β-arrestin recruitment. These results suggest that backbone modification via α → β residue replacement may be a versatile source of agonists with biased GLP-1R activation profiles.
Co-reporter:Dale F. Kreitler; David E. Mortenson; Katrina T. Forest
Journal of the American Chemical Society 2016 Volume 138(Issue 20) pp:6498-6505
Publication Date(Web):May 12, 2016
DOI:10.1021/jacs.6b01454
Synthetic peptides that contain backbone modifications but nevertheless adopt folded structures similar to those of natural polypeptides are of fundamental interest and may provide a basis for biomedical applications. Such molecules can, for example, mimic the ability of natural prototypes to bind to specific target macromolecules but resist degradation by proteases. We have previously shown that oligomers containing mixtures of α- and β-amino acid residues (“α/β-peptides”) can mimic the α-helix secondary structure, and that properly designed α/β-peptides can bind to proteins that evolved to bind to α-helical partners. Here we report fundamental studies that support the long-range goal of extending the α/β approach to tertiary structures. We have evaluated the impact of single α → β modifications on the structure and stability of the small and well-studied villin headpiece subdomain (VHP). The native state of this 35-residue polypeptide contains several α-helical segments packed around a small hydrophobic core. We examined α → β substitution at four solvent-exposed positions, Asn19, Trp23, Gln26 and Lys30. In each case, both the β3 homologue of the natural α residue and a cyclic β residue were evaluated. All α → β3 substitutions caused significant destabilization of the tertiary structure as measured by variable-temperature circular dichroism, although at some of these positions, replacing the β3 residue with a cyclic β residue led to improved stability. Atomic-resolution structures of four VHP analogues were obtained via quasiracemic crystallization. These findings contribute to a fundamental α/β-peptide knowledge-base by confirming that β3-amino acid residues can serve as effective structural mimics of homologous α-amino acid residues within a natural tertiary fold, which should support rational design of functional α/β analogues of natural poly-α-peptides.
Co-reporter:Melissa J. MacDonald, Luke D. Lavis, Donald Hilvert, and Samuel H. Gellman
Organic Letters 2016 Volume 18(Issue 15) pp:3518-3521
Publication Date(Web):July 11, 2016
DOI:10.1021/acs.orglett.6b01279
Efficient hydrolysis of amide bonds has long been a reaction of interest for organic chemists. The rate constants of proteases are unmatched by those of any synthetic catalyst. It has been proposed that a dipeptide containing serine and histidine is an effective catalyst of amide hydrolysis, based on an apparent ability to degrade a protein. The capacity of the Ser-His dipeptide to catalyze the hydrolysis of several discrete ester and amide substrates is investigated using previously described conditions. This dipeptide does not catalyze the hydrolysis of amide or unactivated ester groups in any of the substrates under the conditions evaluated.
Co-reporter:Ross W. Cheloha, Tomoyuki Watanabe, Thomas Dean, Samuel H. Gellman, and Thomas J. Gardella
ACS Chemical Biology 2016 Volume 11(Issue 10) pp:2752
Publication Date(Web):August 17, 2016
DOI:10.1021/acschembio.6b00404
A backbone-modified peptide derived from parathyroid hormone (PTH) is shown to function as an inhibitor and inverse agonist of parathyroid hormone receptor-1 (PTHR1) signaling. This receptor acts to regulate calcium and phosphate homeostasis, as well as bone turnover and development. PTH is a natural agonist of PTHR1, and PTH(1–34) displays full activity relative to the natural 84-residue hormone. PTH(1–34) is used clinically to treat osteoporosis. N-terminally truncated derivatives of PTH(1–34), such as PTH(7–34), are known to bind to PTHR1 without initiating intracellular signaling and can thus act as competitive antagonists of PTH-induced signaling at PTHR1. In some cases, N-terminally truncated PTH derivatives also act as inverse agonists of PTHR1 variants that display pathologically high levels of signaling in the absence of PTH. Many analogues of PTH, however, are rapidly degraded by proteases, which may limit biomedical application. We show that backbone modification via periodic replacement of α-amino acid residues with homologous β-amino acid residues leads to an α/β-PTH(7–34) peptide that retains the antagonist and inverse agonist activities of the prototype α-peptide while exhibiting enhanced stability in the presence of aggressive proteases. These findings highlight the value of backbone-modified peptides derived from PTH as tools for investigating determinants of PTH metabolism and provide guidance for designing therapeutic agents for diseases arising from excessive ligand-dependent or ligand-independent PTHR1 activity.
Co-reporter:Heejun Choi, Saswata Chakraborty, Runhui Liu, Samuel H. Gellman, and James C. Weisshaar
ACS Chemical Biology 2016 Volume 11(Issue 1) pp:113
Publication Date(Web):October 22, 2015
DOI:10.1021/acschembio.5b00547
Synthetic random copolymers based on the nylon-3 (β-peptide) backbone show promise as inexpensive antimicrobial agents resistant to proteolysis. We present a time-resolved observational study of the attack of a particular copolymer MM63:CHx37 on single, live Escherichia coli cells. The composition and chain length of MM63:CHx37 (63% cationic subunits, 37% hydrophobic subunits, 35-subunit average length) were optimized to enhance antibacterial activity while minimizing lysis of human red blood cells. For E. coli cells that export GFP to the periplasm, we obtain alternating phase-contrast and green fluorescence images with a time resolution of 12 s over 60 min following initiation of copolymer flow. Within seconds, cells shrink and exhibit the same plasmolysis spaces that occur following abrupt external osmotic upshift. The osmoprotection machinery attempts to replenish cytoplasmic water, but recovery is interrupted by permeabilization of the cytoplasmic membrane (CM) to GFP. Evidently, the highly cationic copolymer and its counterions rapidly translocate across the outer membrane without permeabilizing it to GFP. The CM permeabilization event is spatially localized. Cells whose CM has been permeabilized never recover growth. The minimum inhibitory concentration (MIC) for cells lacking the osmolyte importer ProP is 4-fold smaller than for normal cells, suggesting that osmoprotection is an important survival strategy. In addition, at the time of CM permeabilization, we observe evidence of oxidative stress. The MIC under anaerobic conditions is at least 8-fold larger than under aerobic conditions, further implicating oxidative damage as an important bacteriostatic effect. Once the copolymer reaches the periplasm, multiple growth-halting mechanisms proceed in parallel.
Co-reporter:Runhui Liu; Xinyu Chen; Shaun P. Falk; Kristyn S. Masters; Bernard Weisblum
Journal of the American Chemical Society 2015 Volume 137(Issue 6) pp:2183-2186
Publication Date(Web):February 4, 2015
DOI:10.1021/ja512567y
Candida albicans is the most common fungal pathogen in humans, and most diseases produced by C. albicans are associated with biofilms. We previously developed nylon-3 polymers with potent activity against planktonic C. albicans and excellent C. albicans versus mammalian cell selectivity. Here we show that these nylon-3 polymers have strong and selective activity against drug-resistant C. albicans in biofilms, as manifested by inhibition of biofilm formation and by killing of C. albicans in mature biofilms. The best nylon-3 polymer (poly-βNM) is superior to the antifungal drug fluconazole for all three strains examined. This polymer is slightly less effective than amphotericin B (AmpB) for two strains, but the polymer is superior against an AmpB-resistant strain.
Co-reporter:Brian F. Fisher; Li Guo; Brian S. Dolinar; Ilia A. Guzei
Journal of the American Chemical Society 2015 Volume 137(Issue 20) pp:6484-6487
Publication Date(Web):May 14, 2015
DOI:10.1021/jacs.5b03382
Structural characterization of new α/γ-peptide foldamers containing the cyclically constrained γ-amino acid I is described. Crystallographic and 2D NMR analysis shows that γ residue I promotes the formation of a 12/10-helical secondary structure in α/γ-peptides. This helix contains two different types of internal H-bond, and the data show that the 12-atom C═O(i) → H–N(i+3) H-bond is more favorable than the 10-atom C═O(i) → H–N(i–1) H-bond. Several foldamer helices featuring topologically distinct H-bonds have been discovered, but our findings are the first to show that such H-bonds may differ in their favorability.
Co-reporter:Zvi Hayouka; Nicole C. Thomas; David E. Mortenson; Kenneth A. Satyshur; Bernard Weisblum; Katrina T. Forest
Journal of the American Chemical Society 2015 Volume 137(Issue 37) pp:11884-11887
Publication Date(Web):September 10, 2015
DOI:10.1021/jacs.5b07206
Quasiracemic crystallography has been used to explore the significance of homochiral and heterochiral associations in a set of host-defense peptide derivatives. The previously reported racemic crystal structure of a magainin 2 derivative displayed a homochiral antiparallel dimer association featuring a “phenylalanine zipper” notable for the dual roles of phenylalanines in mediating dimerization and formation of an exposed hydrophobic swath. This motif is seen as well in two new quasiracemate crystals that contain the d form of the magainin 2 derivative along with an l-peptide in which one Ala has been replaced by a β-amino acid residue. This structural trend supports the hypothesis that the Phe zipper motif has functional significance.
Co-reporter:James W. Checco; Erinna F. Lee; Marco Evangelista; Nerida J. Sleebs; Kelly Rogers; Anne Pettikiriarachchi; Nadia J. Kershaw; Geoffrey A. Eddinger; David G. Belair; Julia L. Wilson; Chelcie H. Eller; Ronald T. Raines; William L. Murphy; Brian J. Smith◆; Samuel H. Gellman;W. Douglas Fairlie
Journal of the American Chemical Society 2015 Volume 137(Issue 35) pp:11365-11375
Publication Date(Web):August 28, 2015
DOI:10.1021/jacs.5b05896
Peptides can be developed as effective antagonists of protein–protein interactions, but conventional peptides (i.e., oligomers of l-α-amino acids) suffer from significant limitations in vivo. Short half-lives due to rapid proteolytic degradation and an inability to cross cell membranes often preclude biological applications of peptides. Oligomers that contain both α- and β-amino acid residues (“α/β-peptides”) manifest decreased susceptibility to proteolytic degradation, and when properly designed these unnatural oligomers can mimic the protein-recognition properties of analogous “α-peptides”. This report documents an extension of the α/β-peptide approach to target intracellular protein–protein interactions. Specifically, we have generated α/β-peptides based on a “stapled” Bim BH3 α-peptide, which contains a hydrocarbon cross-link to enhance α-helix stability. We show that a stapled α/β-peptide can structurally and functionally mimic the parent stapled α-peptide in its ability to enter certain types of cells and block protein–protein interactions associated with apoptotic signaling. However, the α/β-peptide is nearly 100-fold more resistant to proteolysis than is the parent stapled α-peptide. These results show that backbone modification, a strategy that has received relatively little attention in terms of peptide engineering for biomedical applications, can be combined with more commonly deployed peripheral modifications such as side chain cross-linking to produce synergistic benefits.
Co-reporter:Kimberly J. Peterson-Kaufman, Holly S. Haase, Melissa D. Boersma, Erinna F. Lee, W. Douglas Fairlie, and Samuel H. Gellman
ACS Chemical Biology 2015 Volume 10(Issue 7) pp:1667
Publication Date(Web):May 7, 2015
DOI:10.1021/acschembio.5b00109
We report progress toward a general strategy for mimicking the recognition properties of specific α-helices within natural proteins through the use of oligomers that are less susceptible than conventional peptides to proteolysis. The oligomers contain both α- and β-amino acid residues, with the density of the β subunits low enough that an α-helix-like conformation can be adopted but high enough to interfere with protease activity. Previous studies with a different protein-recognition system that suggested ring-constrained β residues can be superior to flexible β residues in terms of maximizing α/β-peptide affinity for a targeted protein surface. Here, we use mimicry of the 18-residue Bim BH3 domain to expand the scope of this strategy. Two significant advances have been achieved. First, we have developed and validated a new ring-constrained β residue that bears an acidic side chain, which complements previously known analogues that are either hydrophobic or basic. Second, we have discovered that placing cyclic β residues at sites that make direct contact with partner proteins can lead to substantial discrimination between structurally homologous binding partners, the proteins Bcl-xL and Mcl-1. Overall, this study helps to establish that α/β-peptides containing ring-preorganized β residues can reliably provide proteolytically resistant ligands for proteins that naturally evolved to recognize α-helical partners.
Co-reporter:Ross W. Cheloha, Jeremy A. Sullivan, Tong Wang, Jordan M. Sand, John Sidney, Alessandro Sette, Mark E. Cook, M. Suresh, and Samuel H. Gellman
ACS Chemical Biology 2015 Volume 10(Issue 3) pp:844
Publication Date(Web):January 5, 2015
DOI:10.1021/cb500888q
Oligomers that contain both α- and β-amino acid residues, or “α/β-peptides”, have emerged as promising mimics of signal-bearing polypeptides that can inhibit or augment natural protein–protein interactions. α/β-Peptides that contain a sufficient proportion of β residues evenly distributed along the sequence can be highly resistant to enzymatic degradation, which is favorable with regard to in vivo applications. Little is known, however, about recognition of α/β-peptides by the immune system. Prior studies have focused almost entirely on examples that contain a single β residue; such α/β-peptides frequently retain the immunological profile of the analogous α-peptide. We have conducted α-peptide vs α/β-peptide comparisons involving higher β residue content, focusing on molecules with αααβ and ααβαααβ backbone repeat patterns. Among analogues of an 18-mer derived from the Bim BH3 domain and an 8-mer derived from secreted phospholipase-2 (sPLA2), we find that recognition by antibodies raised against the prototype α-peptide is suppressed by periodic α → β replacements. Complementary studies reveal that antibodies raised against Bim BH3- or sPLA2-derived α/β-peptides fail to recognize prototype α-peptides displaying identical side chain repertoires. Because polypeptides containing d-α-amino acid residues are of growing interest for biomedical applications, we included the enantiomer of the sPLA2-derived α-peptide in these studies; this d-peptide is fully competent as a hapten, but the resulting antibodies do not cross react with the enantiomeric peptide. Among analogues of the 9-mer CD8+ T-cell viral epitope GP33, we observe that periodic α → β replacements suppress participation in the MHC I + peptide + T-cell receptor ternary complexes that activate cytotoxic T-lymphocytes, due in part to disruption of MHC binding.
Co-reporter:Vanessa M. Kung;Dr. Gabriel Cornilescu; Samuel H. Gellman
Angewandte Chemie International Edition 2015 Volume 54( Issue 48) pp:14336-14339
Publication Date(Web):
DOI:10.1002/anie.201506448
Abstract
We have examined whether parallel β-sheet secondary structure becomes more stable as the number of β-strands increases, via comparisons among peptides designed to adopt two- or three-stranded parallel β-sheet conformations in aqueous solution. Our three-strand design is the first experimental model of a triple-stranded parallel β-sheet. Analysis of the designed peptides by nuclear magnetic resonance (NMR) and circular dichroism (CD) spectroscopy supports the hypothesis that increasing the number of β-strands, from two to three, increases the stability of the parallel β-sheet. We present the first experimental evidence for cooperativity in the folding of a triple-stranded parallel β-sheet, and we show how minimal model systems may enable experimental documentation of characteristic properties, such as CD spectra, of parallel β-sheets.
Co-reporter:Vanessa M. Kung;Dr. Gabriel Cornilescu; Samuel H. Gellman
Angewandte Chemie 2015 Volume 127( Issue 48) pp:14544-14547
Publication Date(Web):
DOI:10.1002/ange.201506448
Abstract
We have examined whether parallel β-sheet secondary structure becomes more stable as the number of β-strands increases, via comparisons among peptides designed to adopt two- or three-stranded parallel β-sheet conformations in aqueous solution. Our three-strand design is the first experimental model of a triple-stranded parallel β-sheet. Analysis of the designed peptides by nuclear magnetic resonance (NMR) and circular dichroism (CD) spectroscopy supports the hypothesis that increasing the number of β-strands, from two to three, increases the stability of the parallel β-sheet. We present the first experimental evidence for cooperativity in the folding of a triple-stranded parallel β-sheet, and we show how minimal model systems may enable experimental documentation of characteristic properties, such as CD spectra, of parallel β-sheets.
Co-reporter:James W. Checco;Dale F. Kreitler;Nicole C. Thomas;David G. Belair;Nicholas J. Rettko;William L. Murphy;Katrina T. Forest
PNAS 2015 Volume 112 (Issue 15 ) pp:4552-4557
Publication Date(Web):2015-04-14
DOI:10.1073/pnas.1420380112
Peptide-based agents derived from well-defined scaffolds offer an alternative to antibodies for selective and high-affinity
recognition of large and topologically complex protein surfaces. Here, we describe a strategy for designing oligomers containing
both α- and β-amino acid residues (“α/β-peptides”) that mimic several peptides derived from the three-helix bundle “Z-domain”
scaffold. We show that α/β-peptides derived from a Z-domain peptide targeting vascular endothelial growth factor (VEGF) can
structurally and functionally mimic the binding surface of the parent peptide while exhibiting significantly decreased susceptibility
to proteolysis. The tightest VEGF-binding α/β-peptide inhibits the VEGF165-induced proliferation of human umbilical vein endothelial cells. We demonstrate the versatility of this strategy by showing
how principles underlying VEGF signaling inhibitors can be rapidly extended to produce Z-domain–mimetic α/β-peptides that
bind to two other protein partners, IgG and tumor necrosis factor-α. Because well-established selection techniques can identify
high-affinity Z-domain derivatives from large DNA-encoded libraries, our findings should enable the design of biostable α/β-peptides
that bind tightly and specifically to diverse targets of biomedical interest. Such reagents would be useful for diagnostic
and therapeutic applications.
Co-reporter:David E. Mortenson;Jay D. Steinkruger;Dale F. Kreitler;Dominic V. Perroni;Gregory P. Sorenson;Lijun Huang;Ritesh Mittal;Hyun Gi Yun;Benjamin R. Travis;Mahesh K. Mahanthappa;Katrina T. Forest
PNAS 2015 112 (43 ) pp:13144-13149
Publication Date(Web):2015-10-27
DOI:10.1073/pnas.1507918112
Interactions between polypeptide chains containing amino acid residues with opposite absolute configurations have long been
a source of interest and speculation, but there is very little structural information for such heterochiral associations.
The need to address this lacuna has grown in recent years because of increasing interest in the use of peptides generated
from d amino acids (d peptides) as specific ligands for natural proteins, e.g., to inhibit deleterious protein–protein interactions. Coiled–coil
interactions, between or among α-helices, represent the most common tertiary and quaternary packing motif in proteins. Heterochiral
coiled–coil interactions were predicted over 50 years ago by Crick, and limited experimental data obtained in solution suggest
that such interactions can indeed occur. To address the dearth of atomic-level structural characterization of heterochiral
helix pairings, we report two independent crystal structures that elucidate coiled-coil packing between l- and d-peptide helices. Both structures resulted from racemic crystallization of a peptide corresponding to the transmembrane segment
of the influenza M2 protein. Networks of canonical knobs-into-holes side-chain packing interactions are observed at each helical
interface. However, the underlying patterns for these heterochiral coiled coils seem to deviate from the heptad sequence repeat
that is characteristic of most homochiral analogs, with an apparent preference for a hendecad repeat pattern.
Co-reporter:David E. Mortenson;Jay D. Steinkruger;Dale F. Kreitler;Dominic V. Perroni;Gregory P. Sorenson;Lijun Huang;Ritesh Mittal;Hyun Gi Yun;Benjamin R. Travis;Mahesh K. Mahanthappa;Katrina T. Forest
PNAS 2015 112 (43 ) pp:13144-13149
Publication Date(Web):2015-10-27
DOI:10.1073/pnas.1507918112
Interactions between polypeptide chains containing amino acid residues with opposite absolute configurations have long been
a source of interest and speculation, but there is very little structural information for such heterochiral associations.
The need to address this lacuna has grown in recent years because of increasing interest in the use of peptides generated
from d amino acids (d peptides) as specific ligands for natural proteins, e.g., to inhibit deleterious protein–protein interactions. Coiled–coil
interactions, between or among α-helices, represent the most common tertiary and quaternary packing motif in proteins. Heterochiral
coiled–coil interactions were predicted over 50 years ago by Crick, and limited experimental data obtained in solution suggest
that such interactions can indeed occur. To address the dearth of atomic-level structural characterization of heterochiral
helix pairings, we report two independent crystal structures that elucidate coiled-coil packing between l- and d-peptide helices. Both structures resulted from racemic crystallization of a peptide corresponding to the transmembrane segment
of the influenza M2 protein. Networks of canonical knobs-into-holes side-chain packing interactions are observed at each helical
interface. However, the underlying patterns for these heterochiral coiled coils seem to deviate from the heptad sequence repeat
that is characteristic of most homochiral analogs, with an apparent preference for a hendecad repeat pattern.
Co-reporter:Michael W. Giuliano ; Stacy J. Maynard ; Aaron M. Almeida ; Li Guo ; Ilia A. Guzei ; Lara C. Spencer
Journal of the American Chemical Society 2014 Volume 136(Issue 42) pp:15046-15053
Publication Date(Web):October 10, 2014
DOI:10.1021/ja5076585
H-bonded helices in conventional peptides (containing exclusively homochiral α-amino acid residues) feature a uniform H-bonding directionality, N-terminal side C═O to C-terminal side NH. In contrast, heterochiral α-peptides can form helices in which the H-bond directionality alternates along the backbone because neighboring amide groups are oriented in opposite directions. Alternating H-bond directions are seen also in helices formed by unnatural peptidic backbones, e.g., those containing β- or γ-amino acid residues. In the present study, we used NMR spectroscopy and crystallography to evaluate the conformational preferences of the novel γ-amino acid (1R,2R,3S)-2-(1-aminopropyl)-cyclohexanecarboxylic acid (APCH), which is constrained by a six-membered ring across its Cα–Cβ bond. These studies were made possible by the development of a stereoselective synthesis of N-protected APCH. APCH strongly enforces the α/γ-peptide 12/10-helical secondary structure, which features alternating H-bond directionality. Thus, APCH residues appear to have a conformational propensity distinct from those of other cyclically constrained γ-amino acid residues.
Co-reporter:Saswata Chakraborty ; Runhui Liu ; Zvi Hayouka ; Xinyu Chen ; Jeffrey Ehrhardt ; Qin Lu ; Eileen Burke ; Yiqing Yang ; Bernard Weisblum ; Gerard C. L. Wong ; Kristyn S. Masters
Journal of the American Chemical Society 2014 Volume 136(Issue 41) pp:14530-14535
Publication Date(Web):October 1, 2014
DOI:10.1021/ja507576a
Host-defense peptides (HDPs) are produced by eukaryotes to defend against bacterial infection, and diverse synthetic polymers have recently been explored as mimics of these natural peptides. HDPs are rich in both hydrophobic and cationic amino acid residues, and most HDP-mimetic polymers have therefore contained binary combinations of hydrophobic and cationic subunits. However, HDP-mimetic polymers rarely duplicate the hydrophobic surface and cationic charge density found among HDPs (Hu, K.; et al. Macromolecules 2013, 46, 1908); the charge and hydrophobicity are generally higher among the polymers. Statistical analysis of HDP sequences (Wang, G.; et al. Nucleic Acids Res. 2009, 37, D933) has revealed that serine (polar but uncharged) is a very common HDP constituent and that glycine is more prevalent among HDPs than among proteins in general. These observations prompted us to prepare and evaluate ternary nylon-3 copolymers that contain a modestly polar but uncharged subunit, either serine-like or glycine-like, along with a hydrophobic subunit and a cationic subunit. Starting from binary hydrophobic–cationic copolymers that were previously shown to be highly active against bacteria but also highly hemolytic, we found that replacing a small proportion of the hydrophobic subunit with either of the polar, uncharged subunits can diminish the hemolytic activity with minimal impact on the antibacterial activity. These results indicate that the incorporation of polar, uncharged subunits may be generally useful for optimizing the biological activity profiles of antimicrobial polymers. In the context of HDP evolution, our findings suggest that there is a selective advantage to retaining polar, uncharged residues in natural antimicrobial peptides.
Co-reporter:Stacy J. Maynard ; Aaron M. Almeida ; Yasuharu Yoshimi
Journal of the American Chemical Society 2014 Volume 136(Issue 47) pp:16683-16688
Publication Date(Web):November 13, 2014
DOI:10.1021/ja510265e
Proteinogenic amino acid residues that promote β-sheet secondary structure are hydrophobic (e.g., Ile or Val) or only moderately polar (e.g., Thr). The design of peptides intended to display β-sheet secondary structure in water typically requires one set of residues to ensure conformational stability and an orthogonal set, with charged side chains, to ensure aqueous solubility and discourage self-association. Here we describe new amino acids that manifest substantial β-sheet propensity, by virtue of β-branching, and also bear an ionizable group in the side chain.
Co-reporter:Runhui Liu ; Xinyu Chen ; Shaun P. Falk ; Brendan P. Mowery ; Amy J. Karlsson ; Bernard Weisblum ; Sean P. Palecek ; Kristyn S. Masters
Journal of the American Chemical Society 2014 Volume 136(Issue 11) pp:4333-4342
Publication Date(Web):March 7, 2014
DOI:10.1021/ja500036r
Fungal infections are a major challenge to human health that is heightened by pathogen resistance to current therapeutic agents. Previously, we were inspired by host-defense peptides to develop nylon-3 polymers (poly-β-peptides) that are toxic toward the fungal pathogen Candida albicans but exert little effect on mammalian cells. Based on subsequent analysis of structure–activity relationships among antifungal nylon-3 polymers, we have now identified readily prepared cationic homopolymers active against strains of C. albicans that are resistant to the antifungal drugs fluconazole and amphotericin B. These nylon-3 polymers are nonhemolytic. In addition, we have identified cationic–hydrophobic copolymers that are highly active against a second fungal pathogen, Cryptococcus neoformans, and moderately active against a third pathogen, Aspergillus fumigatus.
Co-reporter:Runhui Liu ; Xinyu Chen ; Saswata Chakraborty ; Justin J. Lemke ; Zvi Hayouka ; Clara Chow ; Rodney A. Welch ; Bernard Weisblum ; Kristyn S. Masters
Journal of the American Chemical Society 2014 Volume 136(Issue 11) pp:4410-4418
Publication Date(Web):March 7, 2014
DOI:10.1021/ja500367u
Binary nylon-3 copolymers containing cationic and hydrophobic subunits can mimic the biological properties of host-defense peptides, but relationships between composition and activity are not yet well understood for these materials. Hydrophobic subunits in previously studied examples have been limited mostly to cycloalkane-derived structures, with cyclohexyl proving to be particularly promising. The present study evaluates alternative hydrophobic subunits that are isomeric or nearly isomeric with the cyclohexyl example; each has four sp3 carbons in the side chains. The results show that varying the substitution pattern of the hydrophobic subunit leads to relatively small changes in antibacterial activity but causes significant changes in hemolytic activity. We hypothesize that these differences in biological activity profile arise, at least in part, from variations among the conformational propensities of the hydrophobic subunits. The α,α,β,β-tetramethyl unit is optimal among the subunits we have examined, providing copolymers with potent antibacterial activity and excellent prokaryote vs eukaryote selectivity. Bacteria do not readily develop resistance to the new antibacterial nylon-3 copolymers. These findings suggest that variation in subunit conformational properties could be generally valuable in the development of synthetic polymers for biological applications.
Co-reporter:Young-Hee Shin ; David E. Mortenson ; Kenneth A. Satyshur ; Katrina T. Forest
Journal of the American Chemical Society 2013 Volume 135(Issue 22) pp:8149-8152
Publication Date(Web):May 23, 2013
DOI:10.1021/ja403319q
Cyclic constraints have proven to be very effective for preorganizing β-amino acid residues and thereby stabilizing β- and α/β-peptide helices, but little is known about possible preorganization effects among γ residues. Here we assess and compare the impact of cyclic preorganization of β and γ residues in the context of a specific α/β/γ-peptide helix. The results show that β residue preorganization is critical for helix stability but that γ residue preorganization is less important.
Co-reporter:Zvi Hayouka ; Saswata Chakraborty ; Runhui Liu ; Melissa D. Boersma ; Bernard Weisblum
Journal of the American Chemical Society 2013 Volume 135(Issue 32) pp:11748-11751
Publication Date(Web):August 5, 2013
DOI:10.1021/ja406231b
Fmoc-based solid-phase synthesis methodology was used to prepare peptide mixtures containing one type of hydrophobic residue and one type of cationic residue. Each mixture was random in terms of sequence but highly controlled in terms of length. Analysis of the antibacterial and hemolytic properties of these mixtures revealed that selective antibacterial activity can be achieved with heterochiral binary mixtures but not homochiral binary mixture, if the proper amino acid residues are used.
Co-reporter:Runhui Liu ; Xinyu Chen ; Samuel H. Gellman ;Kristyn S. Masters
Journal of the American Chemical Society 2013 Volume 135(Issue 44) pp:16296-16299
Publication Date(Web):October 24, 2013
DOI:10.1021/ja408634a
Substrates that selectively encourage the growth of specific cell types are valuable for the engineering of complex tissues. Some cell-selective peptides have been identified from extracellular matrix proteins; these peptides have proven useful for biomaterials-based approaches to tissue repair or regeneration. However, there are very few examples of synthetic materials that display selectivity in supporting cell growth. We describe nylon-3 polymers that support in vitro culture of endothelial cells but do not support the culture of smooth muscle cells or fibroblasts. These materials may be promising for vascular biomaterials applications.
Co-reporter:Zvi Hayouka ; David E. Mortenson ; Dale F. Kreitler ; Bernard Weisblum ; Katrina T. Forest
Journal of the American Chemical Society 2013 Volume 135(Issue 42) pp:15738-15741
Publication Date(Web):October 8, 2013
DOI:10.1021/ja409082w
High-resolution structure elucidation has been challenging for the large group of host-defense peptides that form helices on or within membranes but do not manifest a strong folding propensity in aqueous solution. Here we report the crystal structure of an analogue of the widely studied host-defense peptide magainin 2. Magainin 2 (S8A, G13A, G18A) is a designed variant that displays enhanced antibacterial activity relative to the natural peptide. The crystal structure of magainin 2 (S8A, G13A, G18A), obtained for the racemic form, features a dimerization mode that has previously been proposed to play a role in the antibacterial activity of magainin 2 and related peptides.
Co-reporter:Runhui Liu ; Xinyu Chen ; Zvi Hayouka ; Saswata Chakraborty ; Shaun P. Falk ; Bernard Weisblum ; Kristyn S. Masters
Journal of the American Chemical Society 2013 Volume 135(Issue 14) pp:5270-5273
Publication Date(Web):April 2, 2013
DOI:10.1021/ja4006404
Host-defense peptides inhibit bacterial growth but show little toxicity toward mammalian cells. A variety of synthetic polymers have been reported to mimic this antibacterial selectivity; however, achieving comparable selectivity for fungi is more difficult because these pathogens are eukaryotes. Here we report nylon-3 polymers based on a novel subunit that display potent antifungal activity (MIC = 3.1 μg/mL for Candida albicans) and favorable selectivity (IC10 > 400 μg/mL for 3T3 fibroblast toxicity; HC10 > 400 μg/mL for hemolysis).
Co-reporter:Pil Seok Chae, Rohini R. Rana, Kamil Gotfryd, Søren G. F. Rasmussen, Andrew C. Kruse, Kyung Ho Cho, Stefano Capaldi, Emil Carlsson, Brian Kobilka, Claus J. Loland, Ulrik Gether, Surajit Banerjee, Bernadette Byrne, John K. Lee and Samuel H. Gellman
Chemical Communications 2013 vol. 49(Issue 23) pp:2287-2289
Publication Date(Web):05 Nov 2012
DOI:10.1039/C2CC36844G
The development of a new class of surfactants for membrane protein manipulation, “GNG amphiphiles”, is reported. These amphiphiles display promising behavior for membrane proteins, as demonstrated recently by the high resolution structure of a sodium-pumping pyrophosphatase reported by Kellosalo et al. (Science, 2012, 337, 473).
Co-reporter:Saswata Chakraborty, Runhui Liu, Justin J. Lemke, Zvi Hayouka, Rodney A. Welch, Bernard Weisblum, Kristyn S. Masters, and Samuel H. Gellman
ACS Macro Letters 2013 Volume 2(Issue 8) pp:753
Publication Date(Web):August 6, 2013
DOI:10.1021/mz400239r
Nylon-3 copolymers containing both hydrophobic and cationic subunits can mimic the activity profile of host-defense peptides, if subunit identity and proportion are carefully selected. These sequence- and stereo-random copolymers inhibit bacterial growth at relatively low concentrations, apparently via disruption of bacterial membranes, but they are relatively nondisruptive toward eukaryotic cell membranes (low hemolytic activity). In all previous examples, the hydrophobic subunits have contained cycloalkyl groups that incorporate the backbone Cα–Cβ bond. Here we have explored the effects of using analogous acyclic hydrophobic subunits. The results indicate that replacing cyclic with acyclic hydrophobic subunits has a modest influence on biological properties. This influence appears to arise from differences in subunit flexibility.
Co-reporter:Soo Hyuk Choi;Monika Ivancic;Ilia A. Guzei
European Journal of Organic Chemistry 2013 Volume 2013( Issue 17) pp:3464-3469
Publication Date(Web):
DOI:10.1002/ejoc.201300118
Abstract
(1R,2S)-2-Aminocyclohexanecarboxylic acid (cis-ACHC) is a preorganized β-amino acid. cis-ACHC favors two conformations that feature gauche conformations about the Cα–Cβ bond with torsion angles of opposite signs. The diastereomeric β-amino acid trans-ACHC has been widely studied as a foldamer building block, but cis-ACHC has received less attention in this regard. We examined the conformational behaviour of three types of oligomer: (1) homooligomers of cis-ACHC, (2) β-peptides in which cis-ACHC and β3h-Ala alternate, and (3) 1:1 α/β-peptides in which cis-ACHC and Ala alternate. Two-dimensional NMR experiments suggest that all three types of oligomer adopt extended conformations rather than folded conformations in solution. Two crystal structures of oligomers that contain cis-ACHC residues, a cis-ACHC dimer and an α/β-peptide tetramer, show extended conformations in which the cis-ACHC residues contain six-membered-ring C=O···H–N hydrogen bonds.
Co-reporter:Michael W. Giuliano, Stacy J. Maynard, Aaron M. Almeida, Andrew G. Reidenbach, Li Guo, Emily C. Ulrich, Ilia A. Guzei, and Samuel H. Gellman
The Journal of Organic Chemistry 2013 Volume 78(Issue 24) pp:12351-12361
Publication Date(Web):December 5, 2013
DOI:10.1021/jo401501g
We report the asymmetric synthesis of the γ-amino acid (1R,2R)-2-aminomethyl-1-cyclopentane carboxylic acid (AMCP) and an evaluation of this residue’s potential to promote secondary structure in α/γ-peptides. Simulated annealing calculations using NMR-derived distance restraints obtained for α/γ-peptides in chloroform reveal that AMCP-containing oligomers are conformationally flexible. However, additional evidence suggests that an internally hydrogen-bonded helical conformation is partially populated in solution. From these data, we propose characteristic NOE patterns for the formation of the α/γ-peptide 12/10-helix and discuss the apparent conformational frustration of AMCP-containing oligomers.
Co-reporter:Holly S. Haase ; Kimberly J. Peterson-Kaufman ; Sheeny K. Lan Levengood ; James W. Checco ; William L. Murphy
Journal of the American Chemical Society 2012 Volume 134(Issue 18) pp:7652-7655
Publication Date(Web):May 1, 2012
DOI:10.1021/ja302469a
Diverse strategies have been explored to mimic the surface displayed by an α-helical segment of a protein, with the goal of creating inhibitors of helix-mediated protein–protein interactions. Many recognition surfaces on proteins, however, are topologically more complex and less regular than a single α-helix. We describe efforts to develop peptidic foldamers that bind to the irregular receptor-recognition surface of vascular endothelial growth factor (VEGF). Our approach begins with a 19-residue α-peptide previously reported by Fairbrother et al. (Biochemistry1998, 37, 17754) to bind to this surface on VEGF. Systematic evaluation of α→β replacements throughout this 19-mer sequence enabled us to identify homologues that contain up to ∼30% β residues, retain significant affinity for VEGF, and display substantial resistance to proteolysis. These α/β-peptides can block VEGF-stimulated proliferation of human umbilical vein endothelial cells.
Co-reporter:Lisa M. Johnson ; David E. Mortenson ; Hyun Gi Yun ; W. Seth Horne ; Thomas J. Ketas ; Min Lu ; John P. Moore
Journal of the American Chemical Society 2012 Volume 134(Issue 17) pp:7317-7320
Publication Date(Web):April 23, 2012
DOI:10.1021/ja302428d
We report a new method for preorganization of α/β-peptide helices, based on the use of a dense array of acidic and basic side chains. Previously we have used cyclically constrained β residues to promote α/β-peptide helicity; here we show that an engineered ion pair array can be comparably effective, as indicated by mimicry of the CHR domain of HIV protein gp41. The new design is effective in biochemical and cell-based infectivity assays; however, the resulting α/β-peptide is susceptible to proteolysis. This susceptibility was addressed via introduction of a few cyclic β residues near the cleavage site, to produce the most stable, effective α/β-peptide gp41 CHR analogue identified. Crystal structures of an α- and α/β-peptide (each involved in a gp41-mimetic helix bundle) that contain the dense acid/base residue array manifest disorder in the ionic side chains, but there is little side-chain disorder in analogous α- and α/β-peptide structures with a sparser ionic side-chain array. These observations suggest that dense arrays of complementary acidic and basic residues can provide conformational stabilization via Coulombic attractions that do not require entropically costly ordering of side chains.
Co-reporter:Jay D. Steinkruger ; Gail J. Bartlett ; Derek N. Woolfson
Journal of the American Chemical Society 2012 Volume 134(Issue 38) pp:15652-15655
Publication Date(Web):September 13, 2012
DOI:10.1021/ja3063088
Pairing preferences in heterodimeric coiled coils are determined by complementarities among side chains that pack against one another at the helix–helix interface. However, relationships between dimer stability and interfacial residue identity are not fully understood. In the context of the “knobs-into-holes” (KIH) packing pattern, one can identify two classes of interactions between side chains from different helices: “lateral”, in which a line connecting the adjacent side chains is perpendicular to the helix axes, and “vertical”, in which the connecting line is parallel to the helix axes. We have previously analyzed vertical interactions in antiparallel coiled coils and found that one type of triad constellation (a′–a–a′) exerts a strong effect on pairing preferences, while the other type of triad (d′–d–d′) has relatively little impact on pairing tendencies. Here, we ask whether vertical interactions (d′–a–d′) influence pairing in parallel coiled-coil dimers. Our results indicate that vertical interactions can exert a substantial impact on pairing specificity, and that the influence of the d′–a–d′ triad depends on the lateral a′ contact within the local KIH motif. Structure-informed bioinformatic analyses of protein sequences reveal trends consistent with the thermodynamic data derived from our experimental model system in suggesting that heterotriads involving Leu and Ile are preferred over homotriads involving Leu and Ile.
Co-reporter:David E. Mortenson ; Kenneth A. Satyshur ; Ilia A. Guzei ; Katrina T. Forest
Journal of the American Chemical Society 2012 Volume 134(Issue 5) pp:2473-2476
Publication Date(Web):January 18, 2012
DOI:10.1021/ja210045s
Quasiracemic crystallization has been used to obtain high-resolution structures of two variants of the villin headpiece subdomain (VHP) that contain a pentafluorophenylalanine (F5Phe) residue in the hydrophobic core. In each case, the crystal contained the variant constructed from l-amino acids and the native sequence constructed from d-amino acids. We were motivated to undertake these studies by reports that racemic proteins crystallize more readily than homochiral forms and the prospect that quasiracemic crystallization would enable us to determine whether a polypeptide containing a noncanonical residue can closely mimic the tertiary structure of the native sequence. The results suggest that quasiracemic crystallization may prove to be generally useful for assessing mimicry of naturally evolved protein folding patterns by polypeptides that contain unnatural side-chain or backbone subunits.
Co-reporter:Jay D. Steinkruger ; Gail J. Bartlett ; Erik B. Hadley ; Lindsay Fay ; Derek N. Woolfson
Journal of the American Chemical Society 2012 Volume 134(Issue 5) pp:2626-2633
Publication Date(Web):January 31, 2012
DOI:10.1021/ja208855x
Elucidating relationships between the amino-acid sequences of proteins and their three-dimensional structures, and uncovering non-covalent interactions that underlie polypeptide folding, are major goals in protein science. One approach toward these goals is to study interactions between selected residues, or among constellations of residues, in small folding motifs. The α-helical coiled coil has served as a platform for such studies because this folding unit is relatively simple in terms of both sequence and structure. Amino acid side chains at the helix–helix interface of a coiled coil participate in so-called “knobs-into-holes” (KIH) packing whereby a side chain (the knob) on one helix inserts into a space (the hole) generated by four side chains on a partner helix. The vast majority of sequence–stability studies on coiled-coil dimers have focused on lateral interactions within these KIH arrangements, for example, between an a position on one helix and an a′ position of the partner in a parallel coiled-coil dimer, or between a--d′ pairs in an antiparallel dimer. More recently, it has been shown that vertical triads (specifically, a′--a--a′ triads) in antiparallel dimers exert a significant impact on pairing preferences. This observation provides impetus for analysis of other complex networks of side-chain interactions at the helix–helix interface. Here, we describe a combination of experimental and bioinformatics studies that show that d′--d--d′ triads have much less impact on pairing preference than do a′--a--a′ triads in a small, designed antiparallel coiled-coil dimer. However, the influence of the d′--d--d′ triad depends on the lateral a′--d interaction. Taken together, these results strengthen the emerging understanding that simple pairwise interactions are not sufficient to describe side-chain interactions and overall stability in antiparallel coiled-coil dimers; higher-order interactions must be considered as well.
Co-reporter:Li Guo, Weicheng Zhang, Ilia A. Guzei, Lara C. Spencer, and Samuel H. Gellman
Organic Letters 2012 Volume 14(Issue 10) pp:2582-2585
Publication Date(Web):May 8, 2012
DOI:10.1021/ol3008815
An asymmetric synthesis of two new diastereomeric γ-amino acids is described. Both molecules contain a cyclohexyl ring to limit conformational flexibility about the Cα–Cβ bond; they differ in having cis vs trans stereochemistry on the ring. Residues derived from the cis γ isomer are shown to support helical secondary structures in α/γ-peptide oligomers.
Co-reporter:Jihua Zhang, Matthew J. Markiewicz, Bernard Weisblum, Shannon S. Stahl, and Samuel H. Gellman
ACS Macro Letters 2012 Volume 1(Issue 6) pp:714
Publication Date(Web):May 23, 2012
DOI:10.1021/mz300172y
A new family of β-lactams is described that enables anionic ring-opening polymerization (AROP) to prepare nylon-3 materials bearing diverse appended functionality, including carboxylic acid, thiol, hydroxyl, and secondary amine groups. Nylon-3 copolymers generated with the new β-lactams are shown to display distinctive self-assembly behavior and biological properties.
Co-reporter:Runhui Liu;Kang Z. Vang;Pamela K. Kreeger;Kristyn S. Masters
Journal of Biomedical Materials Research Part A 2012 Volume 100A( Issue 10) pp:2750-2759
Publication Date(Web):
DOI:10.1002/jbm.a.34211
Abstract
The ability to design biomaterials that interact with biological environments in a predictable manner necessitates an improved understanding of how surface chemistry influences events such as protein adsorption and cell adhesion. In this work, we examined mechanisms governing the interactions between 3T3 fibroblasts and nylon-3 polymers, which have a protein-like polyamide backbone and are highly amenable to tuning of chemical and physical properties. Protein adsorption and cell adhesion to a library of nylon-3 polymers were characterized and analyzed by partial least squares regression. This analysis revealed that specific chemical features of the nylon-3 polymers correlated with the extent of protein adsorption, which, in turn, correlated with cell adhesion in a serum-containing environment. In contrast, in a serum-free environment, cell adhesion could be predicted solely from chemical properties. Enzymatic treatments of 3T3 cells before plating indicated that proteins bound to the cell surface mediated cell-nylon-3 polymer interactions under serum-free conditions, with additional analysis suggesting that cell-associated fibronectin played a dominant role in adhesion in the absence of serum. The mechanistic insight gained from these studies can be used to inform the design of new polymer structures in addition to providing a basis for continued development of nylon-3 copolymers for tissue engineering applications. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A 100A:2750–2759, 2012.
Co-reporter:Runhui Liu, Kristyn S. Masters, and Samuel H. Gellman
Biomacromolecules 2012 Volume 13(Issue 4) pp:
Publication Date(Web):March 28, 2012
DOI:10.1021/bm201847n
Nylon-3 polymers have a polyamide backbone reminiscent of that found in proteins (β- vs α-amino acid residues, respectively), which makes these materials interesting for biological applications. Because of the versatility of the ring-opening polymerization process and the variety of β-lactam starting materials available, the structure of nylon-3 copolymers is highly amenable to alteration. A previous study showed that relatively subtle changes in the structure or ratio of hydrophobic and cationic subunits that comprise these polymers can result in significant changes in the ability of nylon-3-bearing surfaces to support cell adhesion and spreading. In the present study, we have exploited the highly tailorable nature of these polymers to synthesize new versions possessing a wide range of chain lengths, with the intent of optimizing these materials for use as cell-supportive substrates. We find that longer nylon-3 chains lead to better fibroblast attachment on modified surfaces and that at the optimal chain lengths less hydrophobic subunits are superior. The best polymers we identified are comparable to an RGD-containing peptide in supporting fibroblast attachment. The results described here will help to focus future efforts aimed at refining nylon-3 copolymer substrates for specific tissue engineering applications.
Co-reporter:Li Guo, Weicheng Zhang, Ilia A. Guzei, Lara C. Spencer, Samuel H. Gellman
Tetrahedron 2012 68(23) pp: 4413-4417
Publication Date(Web):
DOI:10.1016/j.tet.2012.01.079
Co-reporter:Jihua Zhang, Matthew J. Markiewicz, Brendan P. Mowery, Bernard Weisblum, Shannon S. Stahl, and Samuel H. Gellman
Biomacromolecules 2012 Volume 13(Issue 2) pp:
Publication Date(Web):December 14, 2011
DOI:10.1021/bm2013058
Nylon-3 polymers contain β-amino-acid-derived subunits and can be viewed as higher homologues of poly(α-amino acids). This structural relationship raises the possibility that nylon-3 polymers offer a platform for development of new materials with a variety of biological activities, a prospect that has recently begun to receive experimental support. Nylon-3 homo- and copolymers can be prepared via anionic ring-opening polymerization of β-lactams, and use of an N-acyl-β-lactam as coinitiator in the polymerization reaction allows placement of a specific functional group, borne by the N-acyl-β-lactam, at the N-terminus of each polymer chain. Controlling the unit at the C-termini of nylon-3 polymer chains, however, has been problematic. Here we describe a strategy for specifying C-terminal functionality that is based on the polymerization mechanism. After the anionic ring-opening polymerization is complete, we introduce a new β-lactam, approximately 1 equiv relative to the expected number of polymer chains. Because the polymer chains bear a reactive imide group at their C-termini, this new β-lactam should become attached at this position. If the terminating β-lactam bears a distinctive functional group, that functionality should be affixed to most or all C-termini in the reaction mixture. We use the new technique to compare the impact of N- and C-terminal placement of a critical hydrophobic fragment on the biological activity profile of nylon-3 copolymers. The synthetic advance described here should prove to be generally useful for tailoring the properties of nylon-3 materials.
Co-reporter:Melissa D. Boersma, Holly S. Haase, Kimberly J. Peterson-Kaufman, Erinna F. Lee, Oliver B. Clarke, Peter M. Colman, Brian J. Smith, W. Seth Horne, W. Douglas Fairlie, and Samuel H. Gellman
Journal of the American Chemical Society 2011 Volume 134(Issue 1) pp:315-323
Publication Date(Web):October 31, 2011
DOI:10.1021/ja207148m
Peptidic oligomers that contain both α- and β-amino acid residues, in regular patterns throughout the backbone, are emerging as structural mimics of α-helix-forming conventional peptides (composed exclusively of α-amino acid residues). Here we describe a comprehensive evaluation of diverse α/β-peptide homologues of the Bim BH3 domain in terms of their ability to bind to the BH3-recognition sites on two partner proteins, Bcl-xL and Mcl-1. These proteins are members of the anti-apoptotic Bcl-2 family, and both bind tightly to the Bim BH3 domain itself. All α/β-peptide homologues retain the side-chain sequence of the Bim BH3 domain, but each homologue contains periodic α-residue → β3-residue substitutions. Previous work has shown that the ααβαααβ pattern, which aligns the β3-residues in a ’stripe’ along one side of the helix, can support functional α-helix mimicry, and the results reported here strengthen this conclusion. The present study provides the first evaluation of functional mimicry by ααβ and αααβ patterns, which cause the β3-residues to spiral around the helix periphery. We find that the αααβ pattern can support effective mimicry of the Bim BH3 domain, as manifested by the crystal structure of an α/β-peptide bound to Bcl-xL, affinity for a variety of Bcl-2 family proteins, and induction of apoptotic signaling in mouse embryonic fibroblast extracts. The best αααβ homologue shows substantial protection from proteolytic degradation relative to the Bim BH3 α-peptide.
Co-reporter:Aaron M. Almeida ; Rebecca Li
Journal of the American Chemical Society 2011 Volume 134(Issue 1) pp:75-78
Publication Date(Web):December 8, 2011
DOI:10.1021/ja208856c
Disulfide bonds between Cys residues in adjacent strands of parallel β-sheets are rare among proteins, which suggests that parallel β-sheet structure is not stabilized by such disulfide cross-links. We report experimental results that show, surprisingly, that an interstrand disulfide bond can stabilize parallel β-sheets formed by an autonomously folding peptide in aqueous solution. NMR analysis reveals that parallel β-sheet structure is terminated beyond the disulfide bond, which causes deviation from the extended backbone conformation at one of the Cys residues.
Co-reporter:William C. Pomerantz ; Virany M. Yuwono ; Ryan Drake ; Jeffrey D. Hartgerink ; Nicholas L. Abbott
Journal of the American Chemical Society 2011 Volume 133(Issue 34) pp:13604-13613
Publication Date(Web):August 4, 2011
DOI:10.1021/ja204874h
We have examined the effect of β-peptide modifications on the propensity of these helical molecules to form lyotropic liquid crystalline (LC) phases in water. All of the β-peptides we have examined contain 10 residues. In each case, at least three residues are derived from trans-2-aminocyclohexanecarboxylic acid (ACHC), which strongly promotes folding to a 14-helical conformation. The structural features varied include the number of ACHC residues, the nature and spatial arrangement of charged side chains (cationic vs anionic), and the identity of groups at the β-peptide termini. We found that relatively small changes (e.g., swapping the positions of a cationic and an anionic side chain) could have large effects, such as abrogation of LC phase formation. The trends revealed by sequence–property studies led to the design of LC-forming β-peptides that bear biomolecular recognition groups (biotin or the tripeptide Arg–Gly–Asp). Structural analysis via circular dichroism and cryo-transmission electron microscopy revealed the existence of two different types of self-associated species, globular aggregates and nanofibers. Nanofibers are the predominant assembly formed at concentrations that lead to LC phase formation, and we conclude that these nanofibers are the functional mesogens. Overall, these studies show how the modularity of β-peptide oligomers enables elucidation of the relationship between molecular structure and large-scale self-assembly behavior.
Co-reporter:Lisa M. Johnson ; W. Seth Horne
Journal of the American Chemical Society 2011 Volume 133(Issue 26) pp:10038-10041
Publication Date(Web):June 6, 2011
DOI:10.1021/ja203358t
Infection of cells by HIV depends upon profound structural rearrangements within the trimeric viral protein gp41. Critical to this process is the formation of a six-helix bundle in which a set of three N-terminal heptad repeat (NHR) helices assemble to form a core displaying long grooves that provide docking sites for three C-terminal heptad repeat (CHR) helices. We report experiments designed to discriminate between two alternative hypotheses regarding the source of affinity between individual CHR helices and the complementary groove: (1) affinity is dominated by interactions of a small cluster of side chains at one end of the CHR helix; or (2) affinity depends upon interactions distributed across the long CHR helix. We have employed two complementary experimental designs, and results from both favor the latter hypothesis.
Co-reporter:Tomohisa Sawada
Journal of the American Chemical Society 2011 Volume 133(Issue 19) pp:7336-7339
Publication Date(Web):April 26, 2011
DOI:10.1021/ja202175a
Artificial mimicry of α-helices offers a basis for development of protein–protein interaction antagonists. Here we report a new type of unnatural peptidic backbone, containing α-, β-, and γ-amino acid residues in an αγααβα repeat pattern, for this purpose. This unnatural hexad has the same number of backbone atoms as a heptad of α residues. Two-dimensional NMR data clearly establish the formation of an α-helix-like conformation in aqueous solution. The helix formed by our 12-mer α/β/γ-peptide is considerably more stable than the α-helix formed by an analogous 14-mer α-peptide, presumably because of the preorganized β and γ residues employed.
Co-reporter:Dr. Felix Freire;Aaron M. Almeida;Dr. John D. Fisk;Jay D. Steinkruger ; Samuel H. Gellman
Angewandte Chemie 2011 Volume 123( Issue 37) pp:8894-8897
Publication Date(Web):
DOI:10.1002/ange.201102986
Co-reporter:Dr. Li Guo;Weicheng Zhang;Andrew G. Reidenbach;Michael W. Giuliano;Dr. Ilia A. Guzei;Lara C. Spencer ; Samuel H. Gellman
Angewandte Chemie 2011 Volume 123( Issue 26) pp:5965-5968
Publication Date(Web):
DOI:10.1002/ange.201101301
Co-reporter:Dr. Felix Freire;Aaron M. Almeida;Dr. John D. Fisk;Jay D. Steinkruger ; Samuel H. Gellman
Angewandte Chemie International Edition 2011 Volume 50( Issue 37) pp:8735-8738
Publication Date(Web):
DOI:10.1002/anie.201102986
Co-reporter:Dr. Li Guo;Weicheng Zhang;Andrew G. Reidenbach;Michael W. Giuliano;Dr. Ilia A. Guzei;Lara C. Spencer ; Samuel H. Gellman
Angewandte Chemie International Edition 2011 Volume 50( Issue 26) pp:5843-5846
Publication Date(Web):
DOI:10.1002/anie.201101301
Co-reporter:Dr. Erinna F. Lee;Assoc. Brian J. Smith;Dr. W. Seth Horne;Kelsey N. Mayer;Marco Evangelista; Peter M. Colman; Samuel H. Gellman;Dr. W. Douglas Fairlie
ChemBioChem 2011 Volume 12( Issue 13) pp:
Publication Date(Web):
DOI:10.1002/cbic.201190058
Co-reporter:Dr. Erinna F. Lee;Assoc. Brian J. Smith;Dr. W. Seth Horne;Kelsey N. Mayer;Marco Evangelista; Peter M. Colman; Samuel H. Gellman;Dr. W. Douglas Fairlie
ChemBioChem 2011 Volume 12( Issue 13) pp:2025-2032
Publication Date(Web):
DOI:10.1002/cbic.201100314
Abstract
The crystal structure of a complex between the prosurvival protein Bcl-xL and an α/β-peptide 21-mer is described. The α/β-peptide contains six β-amino acid residues distributed periodically throughout the sequence and adopts an α-helix-like conformation that mimics the bioactive shape of the Puma BH3 domain. The α/β-peptide forms all of the noncovalent contacts that have previously been identified as necessary for recognition of the prosurvival protein by an authentic BH3 domain. Comparison of our α/β-peptide:Bcl-xL structure with structures of complexes between native BH3 domains and Bcl-2 family proteins reveals how subtle adjustments, including variations in helix radius and helix bowing, allow the α/β-peptide to engage Bcl-xL with high affinity. Geometric comparisons of the BH3-mimetic α/β-peptide with α/β-peptides in helix-bundle assemblies provide insight on the conformational plasticity of backbones that contain combinations of α- and β-amino acid residues. The BH3-mimetic α/β-peptide displays prosurvival protein-binding preferences distinct from those of Puma BH3 itself, even though these two oligomers have identical side-chain sequences. Our results suggest origins for this backbone-dependent change in selectivity.
Co-reporter:Shlomit Segman-Magidovich;Myung-ryul Lee;Vladimir Vaiser;Bernd Struth;Dr. Samuel H. Gellman;Dr. Hanna Rapaport
Chemistry - A European Journal 2011 Volume 17( Issue 52) pp:14857-14866
Publication Date(Web):
DOI:10.1002/chem.201101775
Abstract
There is growing interest in the design of molecules that undergo predictable self-assembly. Bioinspired oligomers with well-defined conformational propensities are attractive from this perspective, since they can be constructed from diverse building blocks, and self-assembly can be directed by the identities and sequence of the subunits. Here we describe the structure of monolayers formed at the air–water interface by amphiphilic α/β-peptides with 1:1 alternation of α- and β-amino acid residues along the backbone. Two of the α/β-peptides, one a dianion and the other a dication, were used to determine differences between self-assemblies of the net negatively and positively charged oligomers. Two additional α/β-peptides, both zwitterionic, were designed to favor assembly in a 1:1 molar ratio mixture with parallel orientation of neighboring strands. Monolayers formed by these α/β-peptides at the air–water interface were characterized by surface pressure–area isotherms, grazing incidence X-ray diffraction (GIXD), atomic force microscopy and ATR-FTIR. GIXD data indicate that the α/β-peptide assemblies exhibited diffraction features similar to those of β-sheet-forming α-peptides. The diffraction data allowed the construction of a detailed model of an antiparallel α/β-peptide sheet with a unique pleated structure. One of the α/β-peptide assemblies displayed high stability, unparalleled among previously studied assemblies of α-peptides. ATR-FTIR data suggest that the 1:1 mixture of zwitterionic α/β-peptides assembled in a parallel arrangement resembling that of a typical parallel β-sheet secondary structure formed by α-peptides. This study establishes guidelines for design of amphiphilic α/β-peptides that assemble in a predictable manner at an air–water interface, with control of interstrand orientation through manipulation of Coulombic interactions along the backbone.
Co-reporter:Pil Seok Chae, Ilia A. Guzei and Samuel H. Gellman
Journal of the American Chemical Society 2010 Volume 132(Issue 6) pp:1953-1959
Publication Date(Web):January 22, 2010
DOI:10.1021/ja9085148
Tripod amphiphiles are designed to promote the solubilization and stabilization of intrinsic membrane proteins in aqueous solution; facilitation of crystallization is a long-range goal. Membrane proteins are subjects of extensive interest because of their critical biological roles, but proteins of this type can be difficult to study because of their low solubility in water. The nonionic detergents that are typically used to achieve solubility can have the unintended effect of causing protein denaturation. Tripod amphiphiles differ from conventional detergents in that the lipophilic segment contains a branchpoint, and previous work has shown that this unusual amphiphilic architecture can be advantageous relative to traditional detergent structures. Here, we report the crystal structures of several tripod amphiphiles that contain an N-oxide hydrophilic group. The data suggest that tripods can adapt themselves to a nonpolar surface by altering the hydrophobic appendage that projects toward that surface and their overall orientation relative to that surface. Although it is not possible to draw firm conclusions regarding amphiphile association in solution from crystallographic data, trends observed among the packing patterns reported here suggest design strategies to be implemented in future studies.
Co-reporter:Li Guo ; Aaron M. Almeida ; Weicheng Zhang ; Andrew G. Reidenbach ; Soo Hyuk Choi ; Ilia A. Guzei
Journal of the American Chemical Society 2010 Volume 132(Issue 23) pp:7868-7869
Publication Date(Web):May 24, 2010
DOI:10.1021/ja103233a
We report the first high-resolution structural data for the β/γ-peptide 13-helix (i,i+3 C═O···H—N H-bonds), a secondary structure that is formed by oligomers with a 1:1 alternation of β- and γ-amino acid residues. Our characterization includes both crystallophaphic and 2D NMR data. Previous studies suggested that β/γ-peptides constructed from conformationally flexible residues adopt a different helical secondary structure in solution. Our design features preorganized β- and γ-residues, which strongly promote 13-helical folding by the 1:1 β/γ backbone.
Co-reporter:Michelle T. Dohm ; Brendan P. Mowery ; Ann M. Czyzewski ; Shannon S. Stahl ; Samuel H. Gellman ;Annelise E. Barron
Journal of the American Chemical Society 2010 Volume 132(Issue 23) pp:7957-7967
Publication Date(Web):May 20, 2010
DOI:10.1021/ja909734n
Non-natural oligomers have recently shown promise as functional analogues of lung surfactant proteins B and C (SP-B and SP-C), two helical and amphiphilic proteins that are critical for normal respiration. The generation of non-natural mimics of SP-B and SP-C has previously been restricted to step-by-step, sequence-specific synthesis, which results in discrete oligomers that are intended to manifest specific structural attributes. Here we present an alternative approach to SP-B mimicry that is based on sequence-random copolymers containing cationic and lipophilic subunits. These materials, members of the nylon-3 family, are prepared by ring-opening polymerization of β-lactams. The best of the nylon-3 polymers display promising in vitro surfactant activities in a mixed lipid film. Pulsating bubble surfactometry data indicate that films containing the most surface-active polymers attain adsorptive and dynamic-cycling properties that surpass those of discrete peptides intended to mimic SP-B. Attachment of an N-terminal octadecanoyl unit to the nylon-3 copolymers, inspired by the post-translational modifications found in SP-C, affords further improvements by reducing the percent surface area compression to reach low minimum surface tension. Cytotoxic effects of the copolymers are diminished relative to that of an SP-B-derived peptide and a peptoid-based mimic. The current study provides evidence that sequence-random copolymers can mimic the in vitro surface-active behavior of lung surfactant proteins in a mixed lipid film. These findings raise the possibility that random copolymers might be useful for developing a lung surfactant replacement, which is an attractive prospect given that such polymers are easier to prepare than are sequence-specific oligomers.
Co-reporter:Jay D. Steinkruger ; Derek N. Woolfson
Journal of the American Chemical Society 2010 Volume 132(Issue 22) pp:7586-7588
Publication Date(Web):May 14, 2010
DOI:10.1021/ja100080q
A new strategy for rapid evaluation of sequence−stability relationships in the parallel coiled-coil motif is described. The experimental design relies upon thiol−thioester exchange equilibria, an approach that is particularly well suited to examination of heterodimeric systems. Our model system has been benchmarked by demonstrating that it can quantitatively reproduce previously reported trends in interhelical a−a′ side-chain pairing preferences at the coiled-coil interface. This new tool has been used to explore the role of Coulombic interactions between a core position on one helix and a flanking position on the other helix (a−g′). This type of interhelical contact has received relatively little attention to date. Our results indicate that such interactions can influence coiled-coil partner preferences.
Co-reporter:Pil Seok Chae ; Kamil Gotfryd ; Jennifer Pacyna ; Larry J. W. Miercke ; Søren G. F. Rasmussen ; Rebecca A. Robbins ; Rohini R. Rana ; Claus J. Loland ; Brian Kobilka ; Robert Stroud ; Bernadette Byrne ; Ulrik Gether
Journal of the American Chemical Society 2010 Volume 132(Issue 47) pp:16750-16752
Publication Date(Web):November 4, 2010
DOI:10.1021/ja1072959
We describe a new type of synthetic amphiphile that is intended to support biochemical characterization of intrinsic membrane proteins. Members of this new family displayed favorable behavior with four of five membrane proteins tested, and these amphiphiles formed relatively small micelles.
Co-reporter:Soo Hyuk Choi ; Ilia A. Guzei ; Lara C. Spencer
Journal of the American Chemical Society 2010 Volume 132(Issue 39) pp:13879-13885
Publication Date(Web):September 9, 2010
DOI:10.1021/ja1062532
Helices are the most extensively studied secondary structures formed by β-peptide foldamers. Among the five known β-peptide helices, the 12-helix is particularly interesting because the internal hydrogen bond orientation and macrodipole are analogous to those of α-peptide helices (α-helix and 310-helix). The β-peptide 12-helix is defined by i, i+3 C═O···H−N backbone hydrogen bonds and promoted by β-residues with a five-membered ring constraint. The 12-helical scaffold has been used to generate β-peptides with specific biological functions, for which diverse side chains must be properly placed along the backbone and, upon folding, properly arranged in space. Only two crystal structures of 12-helical β-peptides have previously been reported, both for homooligomers of trans-2-aminocyclopentanecarboxylic acid (ACPC). Here we report five additional crystal structures of 12-helical β-peptides, all containing residues that bear side chains. Four of the crystallized β-peptides include trans-4,4-dimethyl-2-aminocyclopentanecarboxylic acid (dm-ACPC) residues, and the fifth contains a β3-hPhe residue. These five β-peptides adopt fully folded 12-helical conformations in the solid state. The new crystal structures, along with previously reported data, allow a detailed characterization of the 12-helical conformation; average backbone torsion angles of β-residues and helical parameters are derived. These structural parameters are found to be similar to those for i, i+3 C═O···H−N hydrogen-bonded helices formed by other peptide backbones generated from α- and/or β-amino acids. The similarity between the conformational behavior of dm-ACPC and ACPC is consistent with previous NMR-based conclusions that 4,4-disubstituted ACPC derivatives are compatible with 12-helical folding. In addition, our data show how a β3-residue is accommodated in the 12-helix, thus enhancing understanding of the diverse conformational behavior of this flexible class of β-amino acids.
Co-reporter:Shlomit Segman;Myung-ryul Lee;Vladimir Vaiser;SamuelH. Gellman Dr.;Hanna Rapaport Dr.
Angewandte Chemie 2010 Volume 122( Issue 4) pp:728-731
Publication Date(Web):
DOI:10.1002/ange.200904566
Co-reporter:JoshuaL. Price;ErikB. Hadley;JayD. Steinkruger ;SamuelH. Gellman Dr.
Angewandte Chemie 2010 Volume 122( Issue 2) pp:378-381
Publication Date(Web):
DOI:10.1002/ange.200904714
Co-reporter:Shlomit Segman;Myung-ryul Lee;Vladimir Vaiser;SamuelH. Gellman Dr.;Hanna Rapaport Dr.
Angewandte Chemie International Edition 2010 Volume 49( Issue 4) pp:716-719
Publication Date(Web):
DOI:10.1002/anie.200904566
Co-reporter:JoshuaL. Price;ErikB. Hadley;JayD. Steinkruger ;SamuelH. Gellman Dr.
Angewandte Chemie International Edition 2010 Volume 49( Issue 2) pp:368-371
Publication Date(Web):
DOI:10.1002/anie.200904714
Co-reporter:Jihua Zhang, Samuel H. Gellman and Shannon S. Stahl
Macromolecules 2010 Volume 43(Issue 13) pp:5618-5626
Publication Date(Web):June 11, 2010
DOI:10.1021/ma1010809
Nylon-3 copolymers generated via ring-opening polymerization of β-lactams have recently been shown to function as selective antibacterial agents or as chemoattractants that can induce fibroblasts to attach to surfaces. Understanding the molecular basis of these activities and developing improved materials requires knowledge of the relative reactivities of different β-lactams, which influence subunit distribution patterns within polymer chains. The homopolymerization of a cyclooctyl β-lactam (2) in the presence of a strong base and imide co-initiator was studied using both gas chromatography (GC) and in situ infrared (IR) spectroscopy. The rate of this anionic ring-opening polymerization reaction exhibits a first-order dependence on the β-lactam and co-initiator concentrations and a zero-order dependence on the base concentration. Analogous studies of four other β-lactams, bearing various substituents [cyclohexyl (1), cyclododecyl (3), and Boc-protected amino groups (4, 5)], revealed that different monomers exhibit significantly different homopolymerization rates. Binary copolymerizations of four β-lactam pairs (1 + 4, 2 + 3, 2 + 4, and 2 + 5), several of which lead to biologically active amphiphilic copolymers, were investigated by GC. In each of the copolymerizations, except for 2 + 3, the two β-lactams were consumed at different rates, leading to compositional drift within the resulting polymers (i.e., variable subunit distribution along the length of the polymer chains). The copolymerization rates of 2 + 3 and 2 + 4 exhibited a monotonic dependence on the starting β-lactam composition, whereas the copolymerization of 1 + 4 and 2 + 5 was slower than either of the respective β-lactam homopolymerizations. Three methods (Fineman−Ross, Kelen−Tudos, and Mayo−Lewis) were employed to determine the reactivity ratios of these β-lactam pairs at low conversions. This analysis confirms that the copolymers obtained from 1 + 4, 2 + 4, or 2 + 5 are characterized by some extent of compositional drift, while poly(2 + 3) is an ideally random copolymer. These results provide valuable insights pertinent to the molecular structure of amphiphilic nylon-3 copolymers that exhibit bioactivity.
Co-reporter:Nickeisha A. Stephenson, Jiang Zhu, Samuel H. Gellman and Shannon S. Stahl
Journal of the American Chemical Society 2009 Volume 131(Issue 29) pp:10003-10008
Publication Date(Web):July 6, 2009
DOI:10.1021/ja8094262
The carbon−nitrogen bond of carboxamides is extremely stable under most conditions. The present study reveals that simple zirconium− and hafnium−amido complexes are highly efficient catalysts for equilibrium-controlled transamidation reactions between secondary amines and tertiary amides. In a number of cases, transamidation proceeds rapidly at room temperature. We find that these new catalysts are sufficiently active to promote the metathesis of tertiary amides, which arises from successive transamidation cycles. The catalytic activities we observe are unprecedented and represent a substantial step toward a long-range goal of conducting equilibrium-controlled reactions with carboxamides.
Co-reporter:Li Guo ; Yonggui Chi ; Aaron M. Almeida ; Ilia A. Guzei ; Brian K. Parker
Journal of the American Chemical Society 2009 Volume 131(Issue 44) pp:16018-16020
Publication Date(Web):October 20, 2009
DOI:10.1021/ja907233q
A highly stereoselective synthesis of novel cyclically constrained γ-amino acid residues is presented. The key step involves organocatalytic Michael addition of an aldehyde to 1-nitrocyclohexene. After aldehyde reduction, this approach provides optically active β-substituted δ-nitro alcohols (96−99% ee), which can be converted to γ-amino acid residues with a variety of substituents at the α position. We have used these new building blocks to prepare α/γ-peptide foldamers that adopt a specific helical conformation in solution and in the solid state.
Co-reporter:Michael W. Giuliano ; W. Seth Horne
Journal of the American Chemical Society 2009 Volume 131(Issue 29) pp:9860-9861
Publication Date(Web):July 6, 2009
DOI:10.1021/ja8099294
Helix bundles are among the most widely studied tertiary and quaternary structural motifs in proteins. Here we present the crystal structure of an α/β-peptide foldamer that adopts a tetrameric helix-bundle quaternary structure with a hydrophobic core composed solely of β-amino acids. The structure displays features that are unprecedented among all known helix bundles composed of either α-peptides or peptidic foldamers. The tetramer is characterized by an asymmetry of interaction between neighboring helices, and the side-chain packing within the hydrophobic core differs fundamentally from the knobs-into-holes arrangement typical of most helix bundles.
Co-reporter:Felix Freire
Journal of the American Chemical Society 2009 Volume 131(Issue 23) pp:7970-7972
Publication Date(Web):May 20, 2009
DOI:10.1021/ja902210f
Pairs of short peptide strands can be induced to adopt an antiparallel β-sheet secondary structure in aqueous solution via a macrocyclic constraint, as illustrated by many natural and designed peptides. We show that an analogous strategy is successful for creation of small units of parallel β-sheet secondary structure in aqueous solution. Cyclization in this case requires nonpeptide segments for N-to-N and C-to-C interstrand linkage. Surprisingly, we find that only one of these segments needs to be preorganized.
Co-reporter:Amy J. Karlsson, William C. Pomerantz, Keane J. Neilsen, Samuel H. Gellman and Sean P. Palecek
ACS Chemical Biology 2009 Volume 4(Issue 7) pp:567
Publication Date(Web):June 11, 2009
DOI:10.1021/cb900093r
β-Peptides (β-amino acid oligomers) that mimic the amphiphilic, helical, and cationic properties of natural antimicrobial peptides have previously been shown to display antifungal activity against planktonic Candida albicans cells. β-Peptides offer several advantages over conventional peptides composed of α-amino acid residues, including conformational stability, resistance to proteases, and activity at physiological salt concentrations. We examined sequence–activity relationships toward both planktonic C. albicans cells and C. albicans biofilms, and the results suggest a toxicity mechanism involving membrane disruption. A strategy for fluorescently labeling a β-peptide without diminishing antifungal activity was devised; labeled β-peptides penetrated the cell membrane and accumulated in the cytoplasm of both planktonic and biofilm-associated cells. The labeled β-peptide was detected only in metabolically inactive cells, which suggests that β-peptide entry is correlated with cell death. The presence of a β-peptide at a concentration near the minimum inhibitory concentration completely prevented planktonic C. albicans cells from forming a biofilm, suggesting that β-peptides may be useful in preventing fungal colonization and biofilm formation.
Co-reporter:William C. Pomerantz Dr.;Erik B. Hadley Dr.;Charles G. Fry Dr.
ChemBioChem 2009 Volume 10( Issue 13) pp:2177-2181
Publication Date(Web):
DOI:10.1002/cbic.200900380
Co-reporter:ErinnaF. Lee Dr.;JackD. Sadowsky;BrianJ. Smith Dr.;PeterE. Czabotar Dr.;KimberlyJ. Peterson-Kaufman;PeterM. Colman ;SamuelH. Gellman ;W.Douglas Fairlie Dr.
Angewandte Chemie International Edition 2009 Volume 48( Issue 24) pp:4318-4322
Publication Date(Web):
DOI:10.1002/anie.200805761
Co-reporter:ManuelM. Müller;MatthewA. Windsor;WilliamC. Pomerantz Dr.;SamuelH. Gellman Dr.;Donald Hilvert Dr.
Angewandte Chemie International Edition 2009 Volume 48( Issue 5) pp:922-925
Publication Date(Web):
DOI:10.1002/anie.200804996
Co-reporter:ErinnaF. Lee Dr.;JackD. Sadowsky;BrianJ. Smith Dr.;PeterE. Czabotar Dr.;KimberlyJ. Peterson-Kaufman;PeterM. Colman ;SamuelH. Gellman ;W.Douglas Fairlie Dr.
Angewandte Chemie 2009 Volume 121( Issue 24) pp:4382-4386
Publication Date(Web):
DOI:10.1002/ange.200805761
Co-reporter:ManuelM. Müller;MatthewA. Windsor;WilliamC. Pomerantz Dr.;SamuelH. Gellman Dr.;Donald Hilvert Dr.
Angewandte Chemie 2009 Volume 121( Issue 5) pp:940-943
Publication Date(Web):
DOI:10.1002/ange.200804996
Co-reporter:W. Seth Horne;Lisa M. Johnson;Thomas J. Ketas;Per Johan Klasse;Min Lu;John P. Moore
PNAS 2009 Volume 106 (Issue 35 ) pp:14751-14756
Publication Date(Web):2009-09-01
DOI:10.1073/pnas.0902663106
Unnatural oligomers that can mimic protein surfaces offer a potentially useful strategy for blocking biomedically important
protein-protein interactions. Here we evaluate an approach based on combining α- and β-amino acid residues in the context
of a polypeptide sequence from the HIV protein gp41, which represents an excellent testbed because of the wealth of available
structural and biological information. We show that α/β-peptides can mimic structural and functional properties of a critical
gp41 subunit. Physical studies in solution, crystallographic data, and results from cell-fusion and virus-infectivity assays
collectively indicate that the gp41-mimetic α/β-peptides effectively block HIV-cell fusion via a mechanism comparable to that
of gp41-derived α-peptides. An optimized α/β-peptide is far less susceptible to proteolytic degradation than is an analogous
α-peptide. Our findings show how a two-stage design approach, in which sequence-based α→β replacements are followed by site-specific
backbone rigidification, can lead to physical and biological mimicry of a natural biorecognition process.
Co-reporter:W. Seth Horne and Samuel H. Gellman
Accounts of Chemical Research 2008 Volume 41(Issue 10) pp:1399
Publication Date(Web):July 1, 2008
DOI:10.1021/ar800009n
The functions performed by proteins and nucleic acids provide the foundation for life. Chemists have recently begun to ask whether it is possible to design synthetic oligomers that approach the structural and functional complexities of these biopolymers. The study of foldamers, non-natural oligomers displaying discrete folding propensities, has demonstrated that there are several synthetic backbones that exhibit biopolymer-like conformational behavior. Early work in this area focused on oligomers comprised of a single type of monomer subunit, but recent efforts have highlighted the potential of mixed or “heterogeneous” backbones to expand the structural and functional repertoire of foldamers. In this Account, we illustrate the promise of heterogeneous backbone foldamers by focusing on examples containing both α- and β-amino acid residues. Some β-residues bear protein-like side chains, while others have cyclic structures that confer conformational rigidity. The study of heterogeneous backbone foldamers has several advantages over that of their homogeneous backbone counterparts, including access to many new molecular shapes based on variations in the stoichiometries and patterns of the subunit combinations and improved prospects for side chain diversification. Recent efforts to develop α/β-peptide foldamers can be divided into two conceptually distinct classes. The first includes entities prepared using a “block” strategy, in which α-peptide segments and β-peptide segments are combined to form a hybrid oligomer. The second class encompasses designs in which α- and β-amino acid monomers are interspersed in a regular pattern throughout an oligomer sequence. One α/β-peptide helical secondary structure, containing C═O(i)···H−N(i+4) H-bonds analogous to those in the α-helix, has been shown via crystallography to form helix bundle quaternary structures. Desirable biological functions have been elicited from α/β-peptide foldamers. Efforts to mimic naturally occurring host-defense α-peptides have yielded new antimicrobial agents and have led to a reexamination of the long-held views regarding structure−activity relationships among these α-peptides and their analogues. Foldamers offer new platforms for mimicry of the molecular surfaces involved in specific protein−protein recognition events; recent achievements in the preparation of α/β-peptide inhibitors of the protein−protein interactions involved in apoptotic signaling (e.g., between Bcl-xL and pro-apoptotic partners) have revealed the benefits of employing heterogeneous backbones relative to homogeneous backbones for foldamer-based designs. These initial successes in the development of α/β-peptides exhibiting specific biological activities highlight the potential of heterogeneous backbone foldamers for use in biomedical applications and provide guidelines for future studies into new target functions.
Co-reporter:Justin K. Murray, Jack D. Sadowsky, Mark Scalf, Lloyd M. Smith, York Tomita and Samuel H. Gellman
ACS Combinatorial Science 2008 Volume 10(Issue 2) pp:204
Publication Date(Web):February 15, 2008
DOI:10.1021/cc700153z
We describe the use of parallel and split-and-mix library synthesis strategies for exploration of structure−activity relationships among peptidic foldamer ligands for the BH3-recognition cleft of the anti-apoptotic protein Bcl-xL. This effort began with a chimeric (α/β+α)-peptide oligomer (composed of an α/β-peptide segment and an α-peptide segment) that we previously identified to bind tightly to the target cleft on Bcl-xL. The side chains that interact with Bcl-xL were varied in a 1000-member one-bead-one-compound library. Fluorescence polarization (FP) screening identified four new analogues with binding affinities similar to that of the lead compound but no analogues with enhanced affinity. These results suggested that significant improvements in affinity were unlikely in this series. We then used library synthesis to examine backbone variations in the C-terminal α-peptide segment of the lead compound. These studies provided an opportunity for direct comparison of parallel and split-and-mix synthesis formats for foldamer libraries with respect to synthetic variability and assay sensitivity. We found that compounds from both the parallel and one-bead-one-compound libraries could be reliably screened in a competition FP assay without purification of library members. Our findings should facilitate the use of combinatorial library synthesis for exploration of foldamers as inhibitors of protein–protein interactions.
Co-reporter:Kimberly J. Peterson, Jack D. Sadowsky, Elizabeth A. Scheef, Soumen Pal, Katerina D. Kourentzi, Richard C. Willson, Emery H. Bresnick, Nader Sheibani, Samuel H. Gellman
Analytical Biochemistry 2008 Volume 378(Issue 1) pp:8-14
Publication Date(Web):1 July 2008
DOI:10.1016/j.ab.2008.03.043
Vascular endothelial growth factor (VEGF) is a homodimeric proangiogenic protein that induces endothelial cell migration and proliferation primarily through interactions with its major receptors, VEGFR-1 and VEGFR-2. Inhibitors of one or both of these VEGF-receptor interactions could be beneficial as therapeutics for diseases caused by dysfunctional angiogenesis (e.g., cancer). Others have reported small peptides that bind to the VEGF dimer at surface regions that are recognized by the receptors. Here we report the development of a fluorescence polarization assay based on the binding to VEGF of a derivative of one of these peptides that has been labeled with BODIPY-tetramethylrhodamine (BODIPYTMR). This 384-well format assay is tolerant to dimethyl sulfoxide (DMSO, up to 4% [v/v]) and has a Z′ factor of 0.76, making it useful for identifying molecules that associate with the receptor-binding surface of the VEGF dimer.
Co-reporter:Erik B. Hadley;Oliver D. Testa;Derek N. Woolfson
PNAS 2008 Volume 105 (Issue 2 ) pp:530-535
Publication Date(Web):2008-01-15
DOI:10.1073/pnas.0709068105
Reliable predictive rules that relate protein sequence to structure would facilitate postgenome predictive biology and the
engineering and de novo design of peptides and proteins. Through a combination of experiment and analysis of the protein data bank (PDB), we have
deciphered and rationalized new rules for helix–helix interfaces of a common protein-folding and association motif, the antiparallel
dimeric coiled coil. These interfaces are defined by a specific pattern of interactions among largely hydrophobic side chains
often referred to as knobs-into-holes (KIH) packing: a knob from one helix inserts into a hole formed by four residues on
the partner. Previous work has focused on lateral interactions within the KIH motif, for example, between an a position on one helix and a d′ position on the other in an antiparallel coiled coil. We show that vertical interactions within the KIH motif, such as a′-a–a′, are energetically important as well. The experimental and database analyses concur regarding preferred vertical combinations,
which can be rationalized as leading to favorable side-chain interactions that we call constellations. The findings presented here highlight an unanticipated level of complexity in coiled-coil interactions, and our analysis
of a few specific constellations illustrates a general, multipronged approach to addressing this complexity.
Co-reporter:W. Seth Horne;Joshua L. Price
PNAS 2008 Volume 105 (Issue 27 ) pp:9151-9156
Publication Date(Web):2008-07-08
DOI:10.1073/pnas.0801135105
The extent to which polypeptide conformation depends on side-chain composition and sequence has been widely studied, but less
is known about the importance of maintaining an α-amino acid backbone. Here, we examine a series of peptides with backbones
that feature different repeating patterns of α- and β-amino acid residues but an invariant side-chain sequence. In the pure
α-backbone, this sequence corresponds to the previously studied peptide GCN4-pLI, which forms a very stable four-helix bundle
quaternary structure. Physical characterization in solution and crystallographic structure determination show that a variety
of α/β-peptide backbones can adopt sequence-encoded quaternary structures similar to that of the α prototype. There is a loss
in helix bundle stability upon β-residue incorporation; however, stability of the quaternary structure is not a simple function
of β-residue content. We find that cyclically constrained β-amino acid residues can stabilize the folds of α/β-peptide GCN4-pLI
analogues and restore quaternary structure formation to backbones that are predominantly unfolded in the absence of cyclic
residues. Our results show a surprising degree of plasticity in terms of the backbone compositions that can manifest the structural
information encoded in a sequence of amino acid side chains. These findings offer a framework for the design of nonnatural
oligomers that mimic the structural and functional properties of proteins.
Co-reporter:W.Seth Horne Dr.;MelissaD. Boersma;MatthewA. Windsor ;SamuelH. Gellman
Angewandte Chemie International Edition 2008 Volume 47( Issue 15) pp:2853-2856
Publication Date(Web):
DOI:10.1002/anie.200705315
Co-reporter:WilliamC. Pomerantz;ViranyM. Yuwono;ClaireL. Pizzey;JefferyD. Hartgerink ;NicholasL. Abbott ;SamuelH. Gellman
Angewandte Chemie International Edition 2008 Volume 47( Issue 7) pp:1241-1244
Publication Date(Web):
DOI:10.1002/anie.200704372
Co-reporter:W.Seth Horne Dr.;MelissaD. Boersma;MatthewA. Windsor ;SamuelH. Gellman
Angewandte Chemie 2008 Volume 120( Issue 15) pp:2895-2898
Publication Date(Web):
DOI:10.1002/ange.200705315
Co-reporter:
Nature Protocols 2007 2(3) pp:
Publication Date(Web):2007-03-22
DOI:10.1038/nprot.2007.23
The first challenge in microwave-assisted combinatorial chemistry is to apply the necessary amount of microwave irradiation for each reaction step in each reaction vessel. During library preparation, microwave-assisted reactions are often carried out in an 'automated sequential' manner, with reaction mixtures being transferred to and from the reaction chamber by the instrument and irradiated individually, one after another, to control experimental conditions (i.e., temperature) accurately for each sample13. However, the time-saving aspect of microwave synthesis is diminished by sequential irradiation steps, even if each sample is irradiated for only a short time (e.g., 96 samples × 5 min reaction time = 8 h total synthesis time). Parallel synthesis, or performing a reaction in multiple reaction vessels simultaneously, can be used to reduce library synthesis time; however, simultaneous exposure of the entire set of reaction vessels (in our case, the reaction vessels are wells in a plate) to microwave irradiation can lead to quite varied synthetic results across the library as a result of inhomogeneous heating14, 15, 16. If insufficient irradiation is applied, then the reaction will not go to completion, but too much microwave irradiation may cause undesirable side reactions. Efforts to develop a suitable reaction vessel17 for microwave-assisted parallel synthesis have produced complex and expensive technological solutions18, 19, 20, 21, 22.We have synthesized a difficult 96-member β-peptide library using these techniques, demonstrating the use of inexpensive polypropylene filter plates and microwave irradiation in a multimode reactor as a simple and effective method for the rapid preparation of peptide libraries on solid support in acceptable purities for direct biological screening23. The 2.5–25-μmol reaction scale generates sufficient material for subsequent HPLC purification and re-screening for validation of the hits without re-synthesis. The synthetic protocol has been used to produce many different peptide libraries of varying aa composition (both α- and β-aa residues) and length (up to 18 residues) (J.K.M., J.D. Sadowsky, S.H.G., manuscript in preparation). With practice, libraries of oligomers containing up to 10 residues can typically be synthesized in a single day.Step 1: Library planning should be given careful consideration, but preparing the platemap requires only 15 min.Steps 2–11: Preparation for the synthesis requires 2–3 h, including REAGENT SETUP.Steps 12–28: Each deprotection/coupling cycle requires 30 min; the overall synthesis time depends on the length of the peptide.Steps 29–37: The cleavage reaction requires 2 h; concentrating the cleavage solution is highly dependent on the flow rate of the nitrogen, often requiring an overnight period.Steps 38–40: Required time is highly dependent on the HPLC and MS equipment available to the researcher and the degree to which the library is characterized, ranging from a few hours to 1 week.If analysis of the crude library members using HPLC and MS reveals that the product mixtures contain substantial amounts of deletion peptide side products resulting from incomplete coupling reactions, then in the second synthesis consider doing Steps 18–24 two times (i.e., double coupling) for the difficult aa coupling steps. Alternatively, modify Step 23 of the difficult coupling steps by subjecting the reaction mixture to multiple cycles (e.g., three cycles) of microwave irradiation, waiting for approximately 10 min between cycles for the plate to cool to room temperature7. Overall, the reaction solvent can also be modified to facilitate difficult couplings2.With the microwave-assisted parallel peptide synthesis protocol, 96 different hexapeptides can be prepared in less than 24 h (excluding characterization). The peptide product mixtures should contain one major species corresponding to the desired product. The initial purity of the peptide products is typically approximately 60% (Fig. 5) and 50% yield (using UV peak area for a given injection volume and a calibration curve to correlate with peptide concentration and hence amount23, 28) for a hexamer, but this decreases with increasing length of the peptide. This protocol has been applied to peptides containing both α- and β-aa with equivalent results.
Co-reporter:Christen M. Bell;Denis A. Kissounko;Samuel H. Gellman ;Shannon S. Stahl
Angewandte Chemie International Edition 2007 Volume 46(Issue 5) pp:
Publication Date(Web):13 DEC 2006
DOI:10.1002/anie.200603588
Trading places: The metathesis of secondary amides through a transacylation mechanism has been achieved by employing catalytic quantities of an organic imide and a Brønsted base (see scheme). Equilibrium-controlled exchange between various amide pairs is demonstrated for substrates bearing N-alkyl and N-aryl substituents.
Co-reporter:Erik B. Hadley;Alex M. Witek;Felix Freire Dr.;Aaron J. Peoples;Samuel H. Gellman
Angewandte Chemie International Edition 2007 Volume 46(Issue 37) pp:
Publication Date(Web):9 AUG 2007
DOI:10.1002/anie.200702449
No need for references: The conformational stability of β hairpins has been examined by replacing a backbone amide group by a thioester (see picture). The increased flexibility causes a decrease in the β-hairpin stability without changing the structure of the folded conformation and allows the stability of the folded state to be measured under native conditions, without the need for reference compounds for the fully unfolded and folded states.
Co-reporter:Christen M. Bell;Denis A. Kissounko;Samuel H. Gellman ;Shannon S. Stahl
Angewandte Chemie 2007 Volume 119(Issue 5) pp:
Publication Date(Web):13 DEC 2006
DOI:10.1002/ange.200603588
Platztausch: Die Metathese sekundärer Amide über eine Transacylierung gelang durch die Verwendung katalytischer Mengen eines organischen Imids und einer Brønsted-Base (siehe Schema). Der gleichgewichtsgesteuerte Austausch zwischen einer Reihe von Amidpaaren wird für Substrate mit N-Alkyl- und N-Arylsubstituenten gezeigt.
Co-reporter:Erik B. Hadley;Alex M. Witek;Felix Freire Dr.;Aaron J. Peoples;Samuel H. Gellman
Angewandte Chemie 2007 Volume 119(Issue 37) pp:
Publication Date(Web):9 AUG 2007
DOI:10.1002/ange.200702449
Kein Vergleich nötig: Die konformative Stabilität von β-Haarnadelstrukturen beim Austausch einer Amidgruppe im Rückgrat gegen einen Thioester wurde untersucht (siehe Bild). Die höhere Flexibilität senkt die Stabilität der β-Haarnadel, ändert die Struktur der gefalteten Konformation aber nicht; überdies kann die Stabilität des gefalteten Zustandes unter nativen Bedingungen ermittelt werden, ohne dass vollständig entfaltete und gefaltete Bezugsverbindungen benötigt werden.
Co-reporter:Ahlke Hayen Dr.;Margaret A. Schmitt;Felix N. Ngassa;Kathryn A. Thomasson Dr. Dr.
Angewandte Chemie 2004 Volume 116(Issue 4) pp:
Publication Date(Web):14 JAN 2004
DOI:10.1002/ange.200352125
Oligopeptide aus mehr als einem Monomerentyp kommen für den Aufbau neuartiger Foldamere infrage. Dieser Vorschlag stützt sich auf Untersuchungen an Oligomeren aus alternierenden L-α- und cyclischen β-Aminosäuren (siehe Bild), die in Lösung eine helicale Sekundärstruktur aufweisen.
Co-reporter:Ahlke Hayen Dr.;Margaret A. Schmitt;Felix N. Ngassa;Kathryn A. Thomasson Dr. Dr.
Angewandte Chemie International Edition 2004 Volume 43(Issue 4) pp:
Publication Date(Web):14 JAN 2004
DOI:10.1002/anie.200352125
Oligopeptides with heterogeneous backbones, i.e., oligopeptides containing more than one monomer type, may be a fruitful source of new foldamers. This conclusion was suggested by studies on oligomers comprising L-α-amino acid and cyclic β-amino acid residues in a sequentially alternating pattern, which are shown to display helical secondary structure in solution.
Co-reporter:Marina Schinnerl;Justin K. Murray;Joseph M. Langenhan;Samuel H. Gellman
European Journal of Organic Chemistry 2003 Volume 2003(Issue 4) pp:
Publication Date(Web):23 JAN 2003
DOI:10.1002/ejoc.200390112
We report a synthesis of a protected derivative of trans-4-aminopiperidine-3-carboxylic acid (APiC). The route provides either enantiomer. All intermediates are purified by crystallization, and large-scale preparation is therefore possible. An analogous route provides either enantiomer of trans-2-aminocyclohexanecarboxylic acid (ACHC). We have previously shown that β-peptide oligomers containing ACHC adopt a helical conformation defined by 14-membered C=O(i)···H−N(i − 2) hydrogen bonds (“14-helix”). Here we show that APiC residues can be incorporated into the 14-helix. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003)
Co-reporter:Arndt M. Brückner;Margarita Garcia;Andrew Marsh;Samuel H. Gellman;Ulf Diederichsen
European Journal of Organic Chemistry 2003 Volume 2003(Issue 18) pp:
Publication Date(Web):5 SEP 2003
DOI:10.1002/ejoc.200300269
Four novel β-amino acids bearing the canonical nucleobases guanine, cytosine, adenine, and thymine in the side chain, are synthesized starting from Boc-L-aspartic acid 4-benzyl ester. The syntheses are accomplished in six steps by the nucleophilic substitution of (S)-β-(tert-butoxycarbonylamino)-δ-bromopentanoic acid benzyl ester with the corresponding nucleobase derivative as the key step. The guaninyl and cytosinyl β-amino acids were built into β-peptides that were studied by temperature-dependent CD and UV spectroscopy. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003)
Co-reporter:Joseph M. Langenhan;Ilia A. Guzei Dr.
Angewandte Chemie International Edition 2003 Volume 42(Issue 21) pp:
Publication Date(Web):28 MAY 2003
DOI:10.1002/anie.200350932
Diamine-linkedsyn-α,β-dialkyl β-amino acid residues (see picture) have been shown to exhibit parallel sheet secondary structure. Adoption of the parallel hairpin folding pattern is less strongly influenced by linker stereochemistry than for similar parallel hairpins in analogous α-peptide systems.
Co-reporter:Joseph M. Langenhan;Ilia A. Guzei Dr.
Angewandte Chemie 2003 Volume 115(Issue 21) pp:
Publication Date(Web):28 MAY 2003
DOI:10.1002/ange.200350932
Diamin-verbrücktesyn-α,β-Dialkyl-β-aminosäureeinheiten (siehe Bild) bilden parallele Faltblatt-Sekundärstrukturen. Die Konfiguration der verbrückenden Gruppe beeinflusst die Bildung der parallelen Haarnadel-Struktureinheiten in diesem Fall in geringerem Maße als bei analogen α-Peptidsystemen.
Co-reporter:Samuel H. Gellman, Ronald T. Raines
Current Opinion in Chemical Biology 2002 Volume 6(Issue 6) pp:727-728
Publication Date(Web):1 December 2002
DOI:10.1016/S1367-5931(02)00405-2
Co-reporter:Tami L. Raguse;Jonathan R. Lai;Samuel H. Gellman
Helvetica Chimica Acta 2002 Volume 85(Issue 12) pp:4154-4164
Publication Date(Web):2 JAN 2003
DOI:10.1002/hlca.200290001
Oligomers of β-substituted β-amino acids (‘β3-peptides') are known to adopt a helical secondary structure defined by 14-membered ring hydrogen bonds ('14-helix'). Here, we describe a deca-β3-peptide, 1, that does not adopt the 14-helical conformation and that may prefer an alternative secondary structure. β3-Peptide 1 is composed exclusively of residues with side chains that are not branched adjacent to the β-C-atom (β3-hLeu, β3-hLys, and β3-hTyr). In contrast, an analogous β-peptide, 2, containing β3-hVal residues in place of the β3-hLeu residues of 1, adopts a 14-helical conformation in MeOH, according to CD data. These results illustrate the importance of side-chain branching in determining the conformational preferences of β3-peptides.
Co-reporter:
Nature Structural and Molecular Biology 2002 9(6) pp:408 - 410
Publication Date(Web):
DOI:10.1038/nsb0602-408
Co-reporter:Heather E. Stanger;Faisal A. Syud;Juan F. Espinosa;Izabela Giriat;Tom Muir
PNAS 2001 Volume 98 (Issue 21 ) pp:12015-12020
Publication Date(Web):2001-10-09
DOI:10.1073/pnas.211536998
Designed peptides that fold autonomously to specific conformations in aqueous solution are useful for elucidating protein
secondary structural preferences. For example, autonomously folding model systems have been essential for establishing the
relationship between α-helix length and α-helix stability, which would be impossible to probe with α-helices embedded in folded
proteins. Here, we use designed peptides to examine the effect of strand length on antiparallel β-sheet stability. α-Helices
become more stable as they grow longer. Our data show that a two-stranded β-sheet (“β-hairpin”) becomes more stable when the
strands are lengthened from five to seven residues, but that further strand lengthening to nine residues does not lead to
further β-hairpin stabilization for several extension sequences examined. (In one case, all-threonine extension, there may
be an additional stabilization on strand lengthening from seven to nine residues.) These results suggest that there may be
an intrinsic limit to strand length for most sequences in antiparallel β-sheet secondary structure.
Co-reporter:Juan F. Espinosa Dr.;Samuel H. Gellman
Angewandte Chemie 2000 Volume 112(Issue 13) pp:
Publication Date(Web):4 JUL 2000
DOI:10.1002/1521-3757(20000703)112:13<2420::AID-ANGE2420>3.0.CO;2-8
Co-reporter:Lisa M. Johnson ; Stacey Barrick ; Marlies V. Hager ; Amanda McFedries ; Edwin A. Homan ; Mary E. Rabaglia ; Mark P. Keller ; Alan D. Attie ; Alan Saghatelian ; Alessandro Bisello
Journal of the American Chemical Society () pp:
Publication Date(Web):September 5, 2014
DOI:10.1021/ja507168t
Glucagon-like peptide-1 (GLP-1) is a natural agonist for GLP-1R, a G protein-coupled receptor (GPCR) on the surface of pancreatic β cells. GLP-1R agoinsts are attractive for treatment of type 2 diabetes, but GLP-1 itself is rapidly degraded by peptidases in vivo. We describe a design strategy for retaining GLP-1-like activity while engendering prolonged activity in vivo, based on strategic replacement of native α residues with conformationally constrained β-amino acid residues. This backbone-modification approach may be useful for developing stabilized analogues of other peptide hormones.
Co-reporter:Joshua L. Price ; W. Seth Horne
Journal of the American Chemical Society () pp:
Publication Date(Web):August 18, 2010
DOI:10.1021/ja103543s
We report high-resolution crystal structures of six new α/β-peptide foldamers that have a regular α-residue/α-residue/β-residue (ααβ) backbone repeat pattern. All of these foldamers were crystallized from aqueous solution, and all display four-helix bundle quaternary structure in the crystalline state. These oligomers are based on the well-studied 33-residue α-peptide GCN4-pLI, which is an engineered derivative of the dimerization domain of GCN4, a yeast transcription factor. GCN4-pLI forms a stable tetramer in solution and crystallizes as a four-helix bundle (Harbury et al. Science 1993, 262, 1401−1407). Previously we described a foldamer (designated 1 here) that was generated from GCN4-pLI by replacing every third α-amino acid residue with the homologous β3-amino acid residue; this ααβ oligomer retains the side chain sequence of the original α-peptide, but the backbone contains 11 additional CH2 units, which are evenly distributed (Horne et al. Proc. Natl. Acad. Sci. U.S.A. 2008, 105, 9151−9156). Despite the expanded backbone, 1 was found to retain the ability to form a tetrameric quaternary structure in which the individual molecules adopt an α-helix-like conformation. Here we compare nine analogues of 1 that have the same ααβ backbone but in which one or more of the flexible β3-amino acid residues is/are replaced with an analogous cyclic β-residue. The motivation for β3→cyclic replacements is to enhance conformational stability; however, a crystal structure of the one previously reported example (designated 2 here) revealed a “stammer” distortion of the helix-bundle architecture relative to 1. The results reported here suggest that the stammer is a peculiarity of 2, because all six of the new α/β-peptides display undistorted four-helix bundle quaternary structures. More broadly, our results indicate that β3→cyclic replacements are generally well-accommodated in helix-bundle quaternary structure, but that such replacements can be destabilizing in certain instances.
Co-reporter:Pil Seok Chae, Rohini R. Rana, Kamil Gotfryd, Søren G. F. Rasmussen, Andrew C. Kruse, Kyung Ho Cho, Stefano Capaldi, Emil Carlsson, Brian Kobilka, Claus J. Loland, Ulrik Gether, Surajit Banerjee, Bernadette Byrne, John K. Lee and Samuel H. Gellman
Chemical Communications 2013 - vol. 49(Issue 23) pp:NaN2289-2289
Publication Date(Web):2012/11/05
DOI:10.1039/C2CC36844G
The development of a new class of surfactants for membrane protein manipulation, “GNG amphiphiles”, is reported. These amphiphiles display promising behavior for membrane proteins, as demonstrated recently by the high resolution structure of a sodium-pumping pyrophosphatase reported by Kellosalo et al. (Science, 2012, 337, 473).
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