Co-reporter:Shuke Wu, Ji Liu, and Zhi Li
ACS Catalysis August 4, 2017 Volume 7(Issue 8) pp:5225-5225
Publication Date(Web):June 29, 2017
DOI:10.1021/acscatal.7b01464
Biocatalytic anti-Markovnikov alkene hydroamination and hydration were achieved based on two concepts involving enzyme cascades: epoxidation–isomerization–amination for hydroamination and epoxidation–isomerization–reduction for hydration. An Escherichia coli strain coexpressing styrene monooxygenase (SMO), styrene oxide isomerase (SOI), ω-transaminase (CvTA), and alanine dehydrogenase (AlaDH) catalyzed the hydroamination of 12 aryl alkenes to give the corresponding valuable terminal amines in high conversion (many ≥86%) and exclusive anti-Markovnikov selectivity (>99:1). Another E. coli strain coexpressing SMO, SOI, and phenylacetaldehyde reductase (PAR) catalyzed the hydration of 12 aryl alkenes to the corresponding useful terminal alcohols in high conversion (many ≥80%) and very high anti-Markovnikov selectivity (>99:1). Importantly, SOI was discovered for stereoselective isomerization of a chiral epoxide to a chiral aldehyde, providing some insights on enzymatic epoxide rearrangement. Harnessing this stereoselective rearrangement, highly enantioselective anti-Markovnikov hydroamination and hydration were demonstrated to convert α-methylstyrene to the corresponding (S)-amine and (S)-alcohol in 84–81% conversion with 97–92% ee, respectively. The biocatalytic anti-Markovnikov hydroamination and hydration of alkenes, utilizing cheap and nontoxic chemicals (O2, NH3, and glucose) and cells, provide an environmentally friendly, highly selective, and high-yielding synthesis of terminal amines and alcohols.Keywords: alkenes; anti-Markovnikov; biocatalysis; enzyme cascades; hydration; hydroamination; Meinwald rearrangement; whole-cell biotransformation;
Co-reporter:Pengfei Gao;Shuke Wu;Prashant Praveen
Applied Microbiology and Biotechnology 2017 Volume 101( Issue 5) pp:1857-1868
Publication Date(Web):2017/03/01
DOI:10.1007/s00253-016-7954-1
Biotransformation is a green and useful tool for sustainable and selective chemical synthesis. However, it often suffers from the toxicity and inhibition from organic substrates or products. Here, we established a hollow fiber membrane bioreactor (HFMB)-based aqueous/organic biphasic system, for the first time, to enhance the productivity of a cascade biotransformation with strong substrate toxicity and inhibition. The enantioselective trans-dihydroxylation of styrene to (S)-1-phenyl-1,2-ethanediol, catalyzed by Escherichia coli (SSP1) coexpressing styrene monooxygenase and an epoxide hydrolase, was performed in HFMB with organic solvent in the shell side and aqueous cell suspension in the lumen side. Various organic solvents were investigated, and n-hexadecane was found as the best for the HFMB-based biphasic system. Comparing to other reported biphasic systems assisted by HFMB, our system not only shield much of the substrate toxicity but also deflate the product recovery burden in downstream processing as the majority of styrene stayed in organic phase while the diol product mostly remained in the aqueous phase. The established HFMB-based biphasic system enhanced the production titer to 143 mM, being 16-fold higher than the aqueous system and 1.6-fold higher than the traditional dispersive partitioning biphase system. Furthermore, the combination of biphasic system with HFMB prevents the foaming and emulsification, thus reducing the burden in downstream purification. HFMB-based biphasic system could serve as a suitable platform for enhancing the productivity of single-step or cascade biotransformation with toxic substrates to produce useful and valuable chemicals.
Co-reporter:Kaiyuan Tian, Kee Tai, Bryan Jian Wei Chua, Zhi Li
Bioresource Technology 2017 Volume 245, Part B(Volume 245, Part B) pp:
Publication Date(Web):1 December 2017
DOI:10.1016/j.biortech.2017.05.108
•The directed evolution of TLL to enhance methanol tolerance for biodiesel production is reported.•High B-factor amino acids residues were identified for iterative saturation mutagenesis.•The best double mutant TLL-S105C/D27R retained 71% activity after incubation in 75% methanol.•E. coli (TLL-S105C/D27R) achieved 81% FAME yield in the biotransformation of waste grease.•The D27R mutation suggested formation of a new hydrogen bond which increased methanol tolerance.Engineering a methanol tolerant lipase is of great importance in biodiesel production. Here, the first semi-rational method for directed enzyme evolution to enhance methanol tolerance by targeting high B-factor residues for iterative saturation mutagenesis (ISM) is reported. The best double mutant, TLL-S105C/D27R, retained 71% of its original activity after incubation in methanol, showing 30% greater methanol tolerance than TLL. TLL-S105C/D27R also displayed 27% higher activity over TLL. Structure modelling suggested that the increased stability of TLL-S105C/D27R was caused by the formation of a new hydrogen bond which stabilized the protein structure. E. coli (TLL-S105C/D27R)-catalyzed biotransformation of waste grease produced biodiesel in 81% yield in 8 h, showing improvement over the 67% yield for E. coli (TLL), while retaining 92% productivity after 4 cycles of biotransformation of waste grease. The engineered TLL mutant shows high potential for commercial biodiesel production.
Co-reporter:Shuke Wu;Yi Zhou;Daniel Seet
Advanced Synthesis & Catalysis 2017 Volume 359(Issue 12) pp:2132-2141
Publication Date(Web):2017/06/19
DOI:10.1002/adsc.201700416
AbstractGreen and selective oxidation methods are highly desired in chemical synthesis and manufacturing. In this work, we have developed a biocatalytic method for the regio- and stereoselective oxidation of styrene derivatives into arylacetic and (S)-2-arylpropionic acids via a one-pot epoxidation–isomerization–oxidation sequence. This was done via the engineering of Escherichia coli (StyABC-EcALDH) coexpressing styrene monooxygenase (SMO), styrene oxide isomerase (SOI) and aldehyde dehydrogenase (EcALDH) as an active and easily available whole-cell catalyst. Regioselective oxidation of styrene and 11 substituted styrenes using the E. coli cells was performed in a one-pot set-up, producing 12 phenylacetic acids in both high conversion and high yield. Engineering of E. coli (StyABC-ADH9v1) coexpressing SMO, SOI and ADH9v1 (a mutated alcohol dehydrogenase) led to biocatalysts capable of regio- and stereoselective oxidation of α-methylstyrene derivatives to the corresponding chiral acids. One-pot asymmetric synthesis of 4 (S)-2-arylpropionic acids was achieved in good conversion and excellent ee with the E. coli cells. This is a new type of asymmetric alkene oxidation to give chiral acids with no chemical counterpart thus far. The cascade bio-oxidation operates under mild conditions, uses molecular oxygen, exhibits very high regio- and enantioselectivity, and gives high conversion, thus providing a green and efficient method for the synthesis of arylacetic acids and (S)-2-arylpropionic acids directly from easily available styrenes.
Co-reporter:Ji Liu;Bryan Q. W. Pang;Dr. Joseph P. Adams;Dr. Radka Snajdrova; Zhi Li
ChemCatChem 2017 Volume 9(Issue 3) pp:425-431
Publication Date(Web):2017/02/06
DOI:10.1002/cctc.201601446
AbstractThe amine dehydrogenase (AmDH) engineered from the phenylalanine dehydrogenase of Rhodococcus sp. M4 was directly immobilized on magnetic nanoparticles (MNP) from the cell-free extract containing his-tagged AmDH through affinity attachment to give AmDH-MNPs with high yield, enzyme loading efficiency, and specific enzyme loading. AmDH-MNPs showed higher activity and productivity than the free enzyme for the asymmetric reductive amination of 4-phenyl-2-butanone 1 a and phenylacetone 1 b, producing the corresponding amines (R)-2 a,b in 99 % ee and 99 % yield, and with recycling of NADH for up to 3956 times. AmDH-MNPs were easily recycled, retaining 91 % of the original productivity in the third cycle of the reductive amination of 1 a. Coupling of immobilized AmDH and immobilized glucose dehydrogenase (GDH) for the asymmetric reductive amination of 1 a gave (R)-2 a in 99 % ee and 74 % yield, with a total turnover number (TTN) of 2940 for NADH recycling. Both immobilized enzymes showed good recyclability, retaining 81 % productivity in the third reaction cycle. The developed method with coupled immobilized AmDH and immobilized GDH for the asymmetric reductive amination of ketones is useful for the synthesis of enantiopure amines, superior to the use of coupled isolated enzymes with enhanced catalytic performance and reduced enzyme cost through catalyst recycling.
Co-reporter:Yi Zhou;Shuke Wu
Advanced Synthesis & Catalysis 2017 Volume 359(Issue 24) pp:4305-4316
Publication Date(Web):2017/12/19
DOI:10.1002/adsc.201700956
AbstractEnantiopure d-phenylglycine and its derivatives are an important group of chiral amino acids with broad applications in thepharmaceutical industry. However, the existing synthetic methods for d-phenylglycine mainly rely on toxic cyanide chemistry and multistep processes. To provide green and safe alternatives, we envisaged cascade biocatalysis for the one-pot synthesis of d-phenylglycine from racemic mandelic acid, styrene, and biobased l-phenylalanine, respectively. Recombinant Escherichia coli (LZ110) was engineered to coexpress four enzymes to catalyze a 3-step reaction in one pot, transforming mandelic acid (210 mM) to give enantiopure d-phenylglycine in 29.5 g L−1 (195 mM) with 93% conversion. Using the same whole-cell catalyst, twelve other d-phenylglycine derivatives were also produced from the corresponding mandelic acid derivatives in high conversion (58–94%) and very high ee (93–99%). E. coli (LZ116) expressing seven enzymes was constructed for the transformation of styrene to enantiopure d-phenylglycine in 80% conversion via a one-pot 6-step cascade biotransformation. Twelve substituted d-phenylglycines were also produced from the corresponding styrene derivatives in high conversion (45–90%) and very high ee (92–99%) via the same cascade reactions. A nine-enzymeexpressing E. coli (LZ143) was engineered to transform biobased l-phenylalanine to enantiopure d-phenylglycine in 83% conversion via a one-pot 8-step transformation. Preparative biotransformations were also demonstrated. The high-yielding synthetic methods use cheap and green reagents (ammonia, glucose, and/or oxygen), and E. coli whole-cell catalysts, thus providing green and useful alternative methods for manufacturing d-phenylglycine.
Co-reporter:Yi Zhou;Shuke Wu
Advanced Synthesis & Catalysis 2017 Volume 359(Issue 24) pp:4259-4259
Publication Date(Web):2017/12/19
DOI:10.1002/adsc.201701445
The front cover picture, provided by Zhi Li, illustrates the use of enzyme cascades to provide new and green ways for the synthesis of chiral molecules. Here, the one-pot synthesis of d-phenylglycines from racemic mandelic acids, styrene, and biobased l-phenylalanine was achieved by using three recombinant Escherichia coli whole-cell catalysts, respectively. The one-pot synthetic process afforded enantiopure d-phenylglycines in high concentration and high yield. Details of this work can be found in the full paper on pages 4305–4316 (Y. Zhou, S. Wu, Z. Li, Adv. Synth. Catal. 2017, 359, 4305–4316; DOI: 10.1002/adsc.201700956).
Co-reporter:Akbar K. Vahidi, Zunsheng Wang, William S. Y. Wong and Zhi Li
Catalysis Science & Technology 2016 vol. 6(Issue 16) pp:6286-6293
Publication Date(Web):18 May 2016
DOI:10.1039/C6CY00755D
O-Acetylserine sulfhydrylase (OASS) was immobilized for the first time as an active and recyclable biocatalyst for the conversion of O-acetyl-L-serine (OAS) with a nucleophile to the corresponding useful and valuable β-substituted L-α-amino acid. The simple and efficient immobilization was achieved via affinity attachment, without costly enzyme purification, by direct treatment of cell free extracts (CFE) of a recombinant Escherichia coli strain expressing His-tagged OASS isozyme CysK or CysM with Ni-NTA functionalized iron oxide magnetic nanoparticles. The obtained nanobiocatalysts His-CysK-MNPs (85 nm) and His-CysM-MNPs (85 nm) showed high specific enzyme loading (150 and 94 mg of protein per g of MNPs, respectively) and enzyme loading efficiency (80 and 100%, respectively) and catalyzed the conversion of OAS and pyrazole to β-pyrazol-1-yl-L-alanine (β-PA) with the full activity (2.3 and 1.7 U mg−1 of protein, respectively) of the corresponding free enzyme. His-CysK-MNPs and His-CysM-MNPs were easily separated under magnetic field and demonstrated good recyclability with the retaining of 57% and 54% productivity in the 10th cycle, respectively. Pretreatment of the nanobiocatalysts with 50 mM β-PA released some weakly bound proteins and improved catalyst stability and recyclability. The pretreated His-CysK-MNPs catalyzed the conversion of OAS and pyrazole to produce 1080 mM β-PA with 60% overall conversion over 20 reaction cycles and retained 62% productivity in the 20th cycle. These results indicate the efficient synthesis of the useful and valuable β-PA, and the immobilized OASS could be useful for the synthesis of other β-substituted L-α-amino acids.
Co-reporter:Jiandong Zhang, Shuke Wu, Jinchuan Wu, and Zhi Li
ACS Catalysis 2015 Volume 5(Issue 1) pp:51
Publication Date(Web):November 11, 2014
DOI:10.1021/cs5016113
A new type of cascade biocatalysis was developed for one-pot enantioselective conversion of a meso- or racemic epoxide to an α-hydroxy ketone in high ee via an epoxide hydrolase-catalyzed hydrolysis of the epoxide, an alcohol dehydrogenase-catalyzed oxidation of the diol intermediate, and an enzyme-catalyzed cofactor regeneration. In vitro cascade biotransformation of meso-epoxides (cyclopentene oxide 1a, cyclohexene oxide 1b, and cycloheptene oxide 1c) was achieved with cell-free extracts containing recombinant SpEH (epoxide hydrolase from Sphingomonas sp. HXN-200), BDHA (butanediol dehydrogenase from Bacillus subtilis BGSC1A1), and LDH (lactate dehydrogenase form Bacillus subtilis) or NOX (NADH oxidase from Lactobacillus brevis DSM 20054), respectively, giving the corresponding (R)-α-hydroxycyclopentanone 3a, (R)-α-hydroxycyclohexanone 3b, and (R)-α-hydroxycycloheptanone 3c in 98–99% ee and 70–50% conversion with TTN of NAD+-recycling of 5500–26 000. Cascade catalysis with mixed cells of Escherichia coli (SpEH) and E. coli (BDHA-NOX) converted 100–300 mM meso-epoxides 1a–1c to (R)-α-hydroxy ketones 3a–3c in 98–99% ee and 85–57% conversion. Cells of E. coli (SpEH-BDHA-NOX) coexpressing all three enzymes were also proven as good catalysts for the cascade conversion of 100–200 mM meso-epoxides 1a–1c, giving (R)-α-hydroxy ketones 3a–3c in 98–99% ee and 79–52% conversion. The cascade biocatalysis for one-pot synthesis of α-hydroxy ketone in high ee was also successfully demonstrated with a racemic epoxide (1,2,3,4-tetrahydronaphthalene-1,2-oxide 1d) as the substrate. By using two whole-cells based approaches, (R)-α-hydroxytetralone 3d was obtained in 99% ee and 49–40% conversion from 20 to 5 mM racemic epoxide 1d. Preparative cascade biotransformation of cyclohexene oxide 1b gave (R)-α-hydroxycyclohexanone 3b in 98% ee with 70% isolated yield. The developed new type of cascade biocatalysis is enantioselective, green, and often high yielding. The concept might be generally applicable to produce other useful enantiopure α-hydroxy ketones from the corresponding meso- or racemic epoxides by cascade catalysis using appropriate enzymes.Keywords: alcohol dehydrogenase; biocatalysis; cascade catalysis; enantioselective synthesis; epoxide hydrolase; α-hydroxy ketone
Co-reporter:Akbar K. Vahidi, Yi Yang, Thao P. N. Ngo, and Zhi Li
ACS Catalysis 2015 Volume 5(Issue 6) pp:3157
Publication Date(Web):April 20, 2015
DOI:10.1021/acscatal.5b00550
A simple method for immobilizing extracellular enzyme without prepurification of the enzyme was developed. Extracellular His-tagged Thermomyces lanuginosus lipase (His-TLL) was immobilized via affinity by direct treatment of core–shell structured iron oxide magnetic nanoparticles containing long-armed nickel-nitrilotriacetic acid surface groups (Ni-NTA-MNPs) with the cell culture supernatant of Pichia pastoris (h-TLL), giving high enzyme loading efficiency, specific enzyme loading, and specific enzyme activity. The nanobiocatalyst His-TLL-MNPs (80 nm) (5 wt % loading) catalyzed the one-pot conversion of waste grease (24 wt % FFA) to biodiesel with 94% yield and showed excellent recyclability. Ni-NTA-MNPs were easily regenerated from the recycled biocatalyst and reusable for enzyme immobilization. The immobilization method was proven to be general by the immobilization of extracellular His-tagged Candida antarctica lipase B (His-CALB) from the cell culture supernatant of P. pastoris (h-CALB).Keywords: biodiesel; enzyme immobilization; extracellular enzyme; grease; magnetic nanoparticles; Thermomyces lanuginosus lipase
Co-reporter:Li Juan Ye, Hui Hung Toh, Yi Yang, Joseph P. Adams, Radka Snajdrova, and Zhi Li
ACS Catalysis 2015 Volume 5(Issue 2) pp:1119
Publication Date(Web):January 15, 2015
DOI:10.1021/cs501906r
Triple mutant K66Q/S149G/N262C (TM_pheDH) of Rhodococcus phenylalanine dehydrogenase (pheDH) was engineered by directed evolution as the first enzyme for the highly enantioselective reductive amination of phenylacetone 1 and 4-phenyl-2-butanone 3, giving (R)-amphetamine 2 and (R)-1-methyl-3-phenylpropylamine 4 in >98% ee, respectively. The new amine dehydrogenase TM_pheDH with special substrate specificity is a valuable addition to the amine dehydrogenase family with very limited number, for asymmetric reductive amination of ketone, an important reaction in sustainable pharmaceutical manufacturing. Molecular docking provided insight into the role of key mutations of pheDH, being useful for engineering new amine dehydrogenases with higher activity and unique substrate scope.Keywords: amine dehydrogenase; biocatalysis; biotransformation; chiral amine; directed evolution; reductive amination
Co-reporter:Priscilia A. Limadinata, Aitao Li and Zhi Li
Green Chemistry 2015 vol. 17(Issue 2) pp:1194-1203
Publication Date(Web):12 Nov 2014
DOI:10.1039/C4GC01742K
The development of an immobilized enzyme for efficient biocatalysis and catalyst recycling is of great importance in cost-effective and green chemical synthesis, with the hydrolysis of cellulose to glucose for the utilization of lignocellulosic biomass as a prominent and challenging example. We developed a novel concept of engineering temperature-responsive nanobiocatalysts with an upper critical solution temperature (UCST) for efficient catalysis as a soluble catalyst at a temperature >UCST and for easy catalyst separation as an insoluble catalyst at a temperature <UCST. The first polymeric nanoparticles showing an UCST were fabricated in high yield, and the immobilization of cellulase and cellobiase onto the particles afforded the first UCST-biocatalysts, with high specific enzyme loading and enzyme loading efficiency, an UCST of 13–14 °C, and high retention of the initial hydrolysis activity of the free enzyme at 50 °C (>UCST). The hydrolysis of insoluble cellulose, such as filter paper and pre-treated oil palm Empty Fruit Bunch (EFB, a waste biomass), with a mixture of the UCST-nanobiocatalysts containing cellulase and cellobiase at 50 °C reached the same catalytic performance as the free enzymes and gave 97% and 93% glucose yield at 2 wt% of cellulase loading, respectively. These catalytic performances are much better than any other known immobilized cellulases, due to the use of soluble catalysts. The catalysts were easily recovered at 4 °C and recycled to retain 71% and 73% productivity in the 8th and 6th reaction cycles for hydrolyzing filter paper and pre-treated EFB, respectively. The engineered UCST-nanobiocatalysts are potentially useful for the practical green hydrolysis of cellulose to glucose.
Co-reporter:Yi Yang, Yu Tse Chi, Hui Hung Toh and Zhi Li
Chemical Communications 2015 vol. 51(Issue 5) pp:914-917
Publication Date(Web):25 Nov 2014
DOI:10.1039/C4CC08479A
Directed evolution of P450pyr created I83M/I82T mutant for highly regioselective terminal hydroxylation of n-butanol to 1,4-butanediol, representing the first achievement of this hydroxylation reaction by chemical or enzymatic methods and an unique example of evolving a hydroxylase to switch the substrate acceptance from a hydrophobic to hydrophilic compound.
Co-reporter:Shuke Wu, Yongzheng Chen, Yi Xu, Aitao Li, Qisong Xu, Anton Glieder, and Zhi Li
ACS Catalysis 2014 Volume 4(Issue 2) pp:409
Publication Date(Web):December 20, 2013
DOI:10.1021/cs400992z
Cascade biocatalysis via intracellular epoxidation and hydrolysis was developed as a green and efficient method for enantioselective dihydroxylation of aryl olefins to prepare chiral vicinal diols in high ee and high yield. Escherichia coli (SSP1) coexpressing styrene monooxygenase (SMO) and epoxide hydrolase SpEH was developed as a simple and efficient biocatalyst for S-enantioselective dihydroxylation of terminal aryl olefins 1a–15a to give (S)-vicinal diols 1c–15c in high ee (97.5–98.6% for 10 diols; 92.2–93.9% for 3 diols) and high yield (91–99% for 6 diols; 86–88% for 2 diols; 67% for 3 diols). Combining SMO and epoxide hydrolase StEH showing complementary regioselectivity to SpEH as a biocatalyst for the cascade biocatalysis gave rise to R-enantioselective dihydroxylation of aryl olefins, being the first example of this kind of reversing the overall enantioselectivity of cascade biocatalysis. E. coli (SST1) coexpressing SMO and StEH was also engineered as a green and efficient biocatalyst for R-dihydroxylation of terminal aryl olefins 1a–15a to give (R)-vicinal diols 1c–15c in high ee (94.2–98.2% for 7 diols; 84.2–89.9% for 6 diols) and high yield (90–99% for 6 diols; 85–89% for 5 diols; 65% for 1 diol). E. coli (SSP1) and E. coli (SST1) catalyzed the trans-dihydroxylation of trans-aryl olefin 16a and cis-aryl olefin 17a with excellent and complementary stereoselectivity, giving each of the four stereoisomers of 1-phenyl-1,2-propanediol 16c in high ee and de, respectively. Both strains catalyzed the trans-dihydroxylation of aryl cyclic olefins 18a and 19a to afford the same trans-cyclic diols (1R,2R)-18c and (1R,2R)-19c, respectively, in excellent ee and de. This type of cascade biocatalysis provides a tool that is complementary to Sharpless dihydroxylation, accepting cis-alkene and offering enantioselective trans-dihydroxylation.Keywords: biocatalysis; biotransformations; cascade catalysis; dihydroxylation; enantioselectivity; epoxide hydrolase; monooxygenase; vicinal diol
Co-reporter:Pengfei Gao, Aitao Li, Heng Hiang Lee, Daniel I. C. Wang, and Zhi Li
ACS Catalysis 2014 Volume 4(Issue 10) pp:3763
Publication Date(Web):September 11, 2014
DOI:10.1021/cs5010344
Biocatalytic asymmetric sulfoxidation represents a green method to prepare the useful and valuable enantiopure sulfoxides, but this method sometimes suffers from unsatisfied enantioselectivity and low productivity due to substrate and product inhibitions. Here we developed an aqueous/ionic liquid (IL) biphasic system for simultaneously enhancing the enantioselectivity and productivity of P450 monooxygenase-catalyzed asymmetric sulfoxidations of sulfides 1, 3, 5, 7, and 9, as the first example of this kind for a biooxidation. Escherichia coli (P450pyrI83H-GDH) coexpressing P450pyrI83H monooxygenase and glucose dehydrogenase was engineered for the asymmetric sulfoxidations with cofactor recycling, giving higher R-enantioselectivity than any other known P450 monooxygenases and showing high specific activities. The inhibition to the reactions and the toxicity to the cells of the substrates and products were investigated and mostly avoided by using a KP buffer/[P6,6,6,14][NTf2] biphasic reaction system, in which the IL showed excellent biocompatibility to the cells and high solubility to the substrates and products. Sulfoxidations of 1, 3, 5, 7, and 9 with the resting E. coli cells in the biphasic system increased the product concentration from 9.4 to 20 mM for (R)-phenyl methyl sulfoxide 2, from 1.9 to 9.9 mM for (R)-4-fluorophenyl methyl sulfoxide 4, from 5.4 to 16 mM for (R)-ethyl phenyl sulfoxide 6, from 4.2 to 22 mM for (R)-methyl p-tolyl sulfoxide 8, and from 5.7 to 24 mM for (R)-methyl p-methoxyphenyl sulfoxide 10, respectively, and improved the product ee from 85 to 99% for (R)-2, from 80 to 98% for (R)-4, from 88 to 96% for (R)-6, from 35 to 62% for (R)-8, and from 53 to 67% for (R)-10, respectively. The enhancements in enantioselectivity are possibly caused by the low substrate concentrations in the aqueous phase of the biphasic system. Preparative sulfoxidations to produce the useful and valuable sulfoxides (R)-2, (R)-4, and (R)-6 in 99%, 98%, and 96% ee, respectively, were demonstrated.Keywords: biphasic system; enantioselectivity; ionic liquid; P450 monooxygease; productivity; sulfoxidation
Co-reporter:Zillillah, Toh Ann Ngu and Zhi Li
Green Chemistry 2014 vol. 16(Issue 3) pp:1202-1210
Publication Date(Web):14 Oct 2013
DOI:10.1039/C3GC41379A
A novel, active, and recyclable magnetic nano-size solid acid catalyst was developed for the high-yielding transformation of waste grease to biodiesel (fatty acid methyl esters, FAMEs) via simultaneous esterification of free fatty acids (FFAs) and transesterification of triglycerides with methanol in one pot. The core–shell structured magnetic nanoparticles (MNPs) HPW–PGMA–MNPs consist of iron oxide MNPs as the core, poly(glycidyl methacrylate) (PGMA) as the shell, and phosphotungstic acid (HPW) as the surface acid group. They were conveniently prepared in a 93% yield from PGMA–MNPs via phosphonation with Na2HPO4 and subsequent treatment with Na2WO4·2H2O under acidic conditions, allowing for in situ formation of HPW on the particle surface with a high acidity of 1.13 mmol g−1 and a particle size of 90 nm. The catalyst was fully characterized by EDX, FT-IR, FESEM, TEM, and VSM. It demonstrated a much better catalytic performance for the transesterification of triacetin and for the one-pot transformation of grease to biodiesel than commercially available solid acid catalysts such as Amberlyst 15, Purolite CT-275, and zeolite. One-pot transformation of grease (21.3 wt% FFAs) with methanol using HPW–PGMA–MNPs (4 wt%) gave a 98% FAME yield after 24 h, with 96% conversion for the esterification and >98% conversion for the transesterification. HPW–PGMA–MNPs were easily separated from the reaction mixture under a magnetic field and efficiently reused for further cycles of transformation, retaining 95% productivity in the 10th reaction cycle. Thus, the developed magnetic nano-size solid acid catalyst is potentially useful for the green and economic production of biodiesel from waste grease.
Co-reporter:Renliang Huang, Shuke Wu, Aitao Li and Zhi Li
Journal of Materials Chemistry A 2014 vol. 2(Issue 6) pp:1672-1676
Publication Date(Web):02 Dec 2013
DOI:10.1039/C3TA14323F
A novel concept integrating supramolecular self-assembly and electrostatic complexation at an aqueous liquid–liquid interface to synthesize stable peptide–polymer hybrid capsules was developed. The concept was further applied for enzyme immobilization to give stable and active biocatalysts with low enzyme leakage and high encapsulation efficiency, enzyme loading, and recyclability.
Co-reporter:Ji Liu, Jinchuan Wu and Zhi Li
Chemical Communications 2014 vol. 50(Issue 68) pp:9729-9732
Publication Date(Web):04 Jul 2014
DOI:10.1039/C4CC04150J
Enoyl-ACP reductase (FabI) was identified as a non-OYE ‘ene’-reductase for asymmetric reduction of the CC double bond of α, β-unsaturated ketones. Reduction of several 2-alkylidenecyclopentanones with A-FabI and E-FabI gave (R)-2-alkylcyclopentanones in 95–90% and 70–81% ee, respectively. The product ee was improved to 99–98% in high yield by subsequent one-pot biooxidation.
Co-reporter:Aitao Li, Shuke Wu, Joseph P. Adams, Radka Snajdrova and Zhi Li
Chemical Communications 2014 vol. 50(Issue 63) pp:8771-8774
Publication Date(Web):12 Jun 2014
DOI:10.1039/C4CC03491K
P450tol monooxygenase was discovered as a unique and highly enantioselective enzyme for asymmetric epoxidation of some terminal alkenes containing electron-withdrawing groups and benzylic hydroxylation of several ethylbenzenes giving the corresponding useful and valuable products, such as (R)-2- and 3-substituted styrene oxides, (S)-4-substituted styrene oxides, and (S)-benzylic alcohols, in high ee.
Co-reporter:Yi Yang;Dr. Ji Liu ;Dr. Zhi Li
Angewandte Chemie 2014 Volume 126( Issue 12) pp:3184-3188
Publication Date(Web):
DOI:10.1002/ange.201311091
Abstract
Terminal-selective cytochrome P450pyr has been successfully engineered through directed evolution for the subterminal hydroxylation of alkanes with excellent regio- and enantioselectivity. A sensitive colorimetric high-throughput screening (HTS) assay was developed for the measurement of both the regioselectivity and the enantioselectivity of a hydroxylation reaction. By using the HTS assay and iterative saturation mutagenesis, sextuple-mutant P450pyrSM1 was created for the hydroxylation of n-octane (1) to give (S)-2-octanol (2) with 98 % ee and >99 % subterminal selectivity. The engineered P450 is the first enzyme for this type of highly selective alkane hydroxylation, being useful for the CH activation and functionalization of alkanes and the preparation of enantiopure alcohols. Molecular modeling provided structure-based understanding of the fully altered regioselectivity and the excellent enantioselectivity. Another sextuple-mutant P450pyrSM2 catalyzed the hydroxylation of propylbenzene (3) to afford (S)-1-phenyl-2-propanol (4) with 95 % ee and 98 % subterminal selectivity.
Co-reporter:Yi Yang;Dr. Ji Liu ;Dr. Zhi Li
Angewandte Chemie International Edition 2014 Volume 53( Issue 12) pp:3120-3124
Publication Date(Web):
DOI:10.1002/anie.201311091
Abstract
Terminal-selective cytochrome P450pyr has been successfully engineered through directed evolution for the subterminal hydroxylation of alkanes with excellent regio- and enantioselectivity. A sensitive colorimetric high-throughput screening (HTS) assay was developed for the measurement of both the regioselectivity and the enantioselectivity of a hydroxylation reaction. By using the HTS assay and iterative saturation mutagenesis, sextuple-mutant P450pyrSM1 was created for the hydroxylation of n-octane (1) to give (S)-2-octanol (2) with 98 % ee and >99 % subterminal selectivity. The engineered P450 is the first enzyme for this type of highly selective alkane hydroxylation, being useful for the CH activation and functionalization of alkanes and the preparation of enantiopure alcohols. Molecular modeling provided structure-based understanding of the fully altered regioselectivity and the excellent enantioselectivity. Another sextuple-mutant P450pyrSM2 catalyzed the hydroxylation of propylbenzene (3) to afford (S)-1-phenyl-2-propanol (4) with 95 % ee and 98 % subterminal selectivity.
Co-reporter:Yi Yang;Dr. Ji Liu ;Dr. Zhi Li
Angewandte Chemie 2014 Volume 126( Issue 12) pp:
Publication Date(Web):
DOI:10.1002/ange.201401084
Co-reporter:Yi Yang;Dr. Ji Liu ;Dr. Zhi Li
Angewandte Chemie International Edition 2014 Volume 53( Issue 12) pp:
Publication Date(Web):
DOI:10.1002/anie.201401084
Co-reporter:Ji Liu and Zhi Li
ACS Catalysis 2013 Volume 3(Issue 5) pp:908
Publication Date(Web):April 8, 2013
DOI:10.1021/cs400101v
The first cascade biotransformation involving enantioselective reduction of a C═C double bond, Baeyer–Villiger oxidation, and lactone hydrolysis was developed as a green and sustainable tool for synthesizing enantiopure δ-lactones. One-pot cascade biotransformations were achieved with Acinetobacter sp. RS1 containing a novel enantioselective reductase and an enantioselective lactone hydrolase and Escherichia coli coexpressing cyclohexanone monooxygenase and glucose dehydrogenase, converting easily available 2-alkylidenecyclopentanones 1–2 into the corresponding valuable flavors and fragrances (R)-δ-lactones 5–6 in high ee. The one-pot synthesis is better than the reported two-step preparation. This concept is useful in developing other redox cascades with the substrates containing C═C double bond.Keywords: Baeyer−Villiger oxidation; bioreduction; cascade biotransformation; enantioselective synthesis; lactone hydrolysis; δ-lactone
Co-reporter:Shuke Wu, Aitao Li, Yit Siang Chin, and Zhi Li
ACS Catalysis 2013 Volume 3(Issue 4) pp:752
Publication Date(Web):March 4, 2013
DOI:10.1021/cs300804v
A unique epoxide hydrolase (SpEH) from Sphingomonas sp. HXN-200 was identified and cloned based on genome sequencing and expressed in Escherichia coli. The engineered E. coli (SpEH) showed the same selectivity and substrate specificity as the wild type strain and 172 times higher activity than Sphingomonas sp. HXN-200 for the hydrolysis of styrene oxide 1. Hydrolysis of racemic styrene oxide 1, substituted styrene oxides 3, 5–7, and N-phenoxycarbonyl-3,4-epoxypiperidine 8 (200–100 mM) with resting cells of E. coli (SpEH) gave (S)-epoxides 1, 3, 5–7 and (−)-8 in 98.0–99.5% enantiomeric excess (ee) and 37.6–46.5% yield. Hydrolysis of cyclopentene oxide 9, cyclohexene oxide 10, and N-benzyloxycarbonyl-3,4-epoxypyrrolidine 11 (100 mM) afforded the corresponding (R, R)-vicinal trans-diols 12–14 in 86–93% ee and 90–99% yield. The ee of (1R, 2R)-cyclohexane-1,2-diol 13 was improved to 99% by simple crystallization. These biotransformations showed high specific activity (0.28–4.3 U/mg cdw), product concentration, product/cells ratio, and cell-based productivity. Hydrolysis at even higher substrate concentration was also achieved: (S)-1 was obtained in 430 mM (51 g/Lorg) and 43% yield; (1R, 2R)-13 was obtained in 500 mM (58 g/L) and >99% yield. Gram-scale preparation of epoxides (S)-1, (S)-3, (S)-6 and diols (1R, 2R)-12, (1R, 2R)-13, (3R, 4R)-14 were also demonstrated. E. coli (SpEH) cells showed the highest enantioselectivity to produce (S)-1 (E of 39) among all known EHs in the form of whole cells or free enzymes and the highest enantioselectivities to produce (S)-3, 5, 6, 7, (−)-8, and (R, R)-14 (E of 36, 35, 28, 57, 22, and 28) among all known EHs. The easily available and highly active E. coli (SpEH) cells are the best biocatalysts known thus far for the practical preparation of these useful and valuable enantiopure epoxides and vicinal diols via hydrolysis.Keywords: biocatalysis; enantioselective hydrolysis; epoxide; epoxide hydrolase; high product concentration; vicinal diol; whole-cell biotransformation
Co-reporter:Aitao Li, Ji Liu, Son Q. Pham and Zhi Li
Chemical Communications 2013 vol. 49(Issue 98) pp:11572-11574
Publication Date(Web):17 Oct 2013
DOI:10.1039/C3CC46675B
A triple mutant of P450pyr monooxygenase (P450pyrTM) catalysed the epoxidation of several para-substituted styrenes as the first enzyme showing high (R)-enantioselectivity and high conversion, demonstrated a broad substrate range, and showed high enantioselectivity for the epoxidation of an unconjugated 1,1-disubstituted alkene, 2-methyl-3-phenyl-1-propene, and a cyclic alkene, N-phenoxycarbonyl-1,2,5,6-tetrahydropyridine, respectively.
Co-reporter:Jiong Zhang;Tingting Xu
Advanced Synthesis & Catalysis 2013 Volume 355( Issue 16) pp:3147-3153
Publication Date(Web):
DOI:10.1002/adsc.201300301
Co-reporter:Lidan Ye;Mohammad Sufian Bin Hudari
Applied Microbiology and Biotechnology 2013 Volume 97( Issue 11) pp:4831-4838
Publication Date(Web):2013 June
DOI:10.1007/s00253-013-4788-y
Cost-effective conversion of lignocellulose hydrolysate to optically pure lactic acid is commercially attractive but very challenging. Bacillus coagulans JI12 was isolated from natural environment and used to produce L-lactic acid (optical purity > 99.5 %) from lignocellulose sugars and acid hydrolysate of oil palm empty fruit bunch (EFB) at 50 °C and pH 6.0 without sterilization of the medium. In fed-batch fermentation with 85 g/L initial xylose and 55 g/L xylose added after 7.5 h, 137.5 g/L lactic acid was produced with a yield of 98 % and a productivity of 4.4 g/L h. In batch fermentation of a sugar mixture containing 8.5 % xylose, 1 % glucose, and 1 % L-arabinose, the lactic acid yield and productivity reached 98 % and 4.8 g/L h, respectively. When EFB hydrolysate was used, 59.2 g/L of lactic acid was produced within 9.5 h at a yield of 97 % and a productivity of 6.2 g/L h, which are the highest among those ever reported from lignocellulose hydrolysates. These results indicate that B. coagulans JI12 is a promising strain for industrial production of L-lactic acid from lignocellulose hydrolysate.
Co-reporter:Zillillah, Guowei Tan and Zhi Li
Green Chemistry 2012 vol. 14(Issue 11) pp:3077-3086
Publication Date(Web):24 Aug 2012
DOI:10.1039/C2GC35779H
Magnetic nano-size solid acid catalysts were prepared, characterized, and examined, for the first time, for the esterification of free fatty acid (FFA) to develop a green and efficient pretreatment step for producing biodiesel (fatty acid methyl ester, FAME) from waste grease containing FFA. PGMA-cat 4, consisting of a core of iron oxide magnetic nanoparticles (MNPs), a poly(glycidyl methacrylate) (PGMA) shell, and sulfonic acid groups on the surface, was synthesized in 98% yield from the corresponding core–shell structured PGMA–MNPs containing epoxy surface groups by gentle sulfonation with Na2SO3, possessing a mean size of 90 nm, high acid capacity of 2.3 mmol H+ g−1, and excellent superparamagnetic properties. Esterification of FFA (16 wt%) in grease with methanol using this catalyst (4 wt%) gave 96% conversion of FFA to FAME within 2 h. PGMA-cat 4 was easily separated under a magnetic field and showed no loss of productivity during 10 cycles. In comparison, PS-cat 12, consisting of a core of iron oxide MNPs, a polystyrene shell, and benzenesulfonic acids on the surface, was active but possessed no recyclability; Si-cat 14, consisting of a core of iron oxide MNPs, a silica shell, and sulfonic acids as surface groups, showed lower activity and poor recycling performance. The catalytic performance of PGMA-cat 4 was also better than those of the micro-size counterpart such as PGMA-cat (m) 15 and large-size solid acid catalysts such as Amberlyst 15. Thus, a novel, active, stable, and recyclable magnetic nano-size solid acid catalyst was successfully developed for the green and efficient esterification of FFA in grease as a pretreatment step to produce biodiesel from waste grease.
Co-reporter:Liang Xue, Shiyao Dai and Zhi Li
Journal of Materials Chemistry A 2012 vol. 22(Issue 15) pp:7403-7411
Publication Date(Web):08 Mar 2012
DOI:10.1039/C2JM15918J
Elastic shape-memory polymers as self-expendable polymeric drug-eluting stents are developed for the first time, with good mechanical properties, fast self-expansion, and sustained drug release. Novel star block co-polymer PCTOPDs containing hyperbranched poly(ε-caprolactone) (PCL) switching segment and poly(2-oxepane-1,5-dione) (POPD) hard segment were synthesized in high yield and characterized by NMR, GPC, DSC, tensile test and cyclic thermomechanical tensile test. PCTOPD containing 15–27 wt% POPD (PCTOPD-15–27) are found to be non-cytotoxic thermoplastic elastomers (Tm of 39–40 and 120–129 °C, εb of 908–1060%, σm of 12–20 MPa, and E of 60–91 MPa) with good shape-memory properties at 40 °C (Rf of 95–97%, Rr of 97–99%, and shape recovery time of 35 s). The stent made from PCTOPD-27 gives nearly full self-expansion at 37 °C within 45 s. The collapse pressure at a compressive strain of 30% is 1.7 bar. A stent containing 3.0 wt% paclitaxel releases 42% drug linearly in the first 9 days and 67% drug in 30 days with a slower but nearly linear release for the period of 10–30 days. The developed SMP-based drug-eluting stents might be useful in biomedical application such as the treatment of coronary artery disease.
Co-reporter:Son Quang Pham, Guillaume Pompidor, Ji Liu, Xiao-Dan Li and Zhi Li
Chemical Communications 2012 vol. 48(Issue 38) pp:4618-4620
Publication Date(Web):05 Mar 2012
DOI:10.1039/C2CC30779K
Directed evolution of a monooxygenase to achieve very high enantioselectivity for hydroxylation at non-activated carbon atoms is demonstrated for the first time, where a triple mutant of P450pyr hydroxylase is obtained via determination of enzyme structure, iterative saturation mutagenesis, and high-throughput screening with a MS-based ee assay to increase the product ee from 53% to 98% for the hydroxylation of N-benzyl pyrrolidine to (S)-N-benzyl 3-hydroxypyrrolidine.
Co-reporter:Thao P. N. Ngo, Wei Zhang, Wen Wang and Zhi Li
Chemical Communications 2012 vol. 48(Issue 38) pp:4585-4587
Publication Date(Web):08 Mar 2012
DOI:10.1039/C2CC30953J
Reversible clusters of nanobiocatalysts are developed via non-covalent interaction among enzyme-bound iron oxide magnetic nanoparticles. Dissociation of the clusters by shaking during biotransformation enables high catalytic performance, and re-clustering by stopping shaking after reaction allows for easy magnetic separation. The novel concept is demonstrated with alcohol dehydrogenase RDR for the enantioselective reduction of 7-methoxy-2-tetralone.
Co-reporter:Xin Jia, Yi Xu, and Zhi Li
ACS Catalysis 2011 Volume 1(Issue 6) pp:591
Publication Date(Web):April 18, 2011
DOI:10.1021/cs200099k
A simple and green method for preparing several enantiopure 1,2-diols was developed, for the first time, via regio- and stereoselective concurrent oxidations of the racemates with microbial cells. Sphingomonas sp. HXN-200 was found to catalyze the regio- and stereoselective oxidations of 3-O-benzylglycerol 1 to the corresponding α-hydroxy aldehyde 5 and then to the α-hydroxy carboxylic acid 6. Concurrent biooxidations of racemic 3-O-benzylglycerol 1 with resting cells of Sphingomonas sp. HXN-200 gave (S)-1 in 99.2% enantiomeric excess (ee) and 32% yield. Similar biooxidations of racemic 1-(4-chlorophenyl)-1,2-ethanediol 2, 1-(4-methylphenyl)-1,2-ethanediol 3, and phenyl-1,2-ethanediol 4 gave (R)-2 in 98.4% ee and 48% yield, (R)-3 in 99.6% ee and 45% yield, and (R)-4 in 98.7% ee and 36% yield, respectively. These represent the best results known thus far for the enzymatic syntheses of the useful and valuable diols (S)-1 and (R)-2–4.Keywords: 2-diols; concurrent oxidations; enantiopure 1; enantioselective synthesis; enzyme catalysis; Sphingomonas sp. HXN-200; tandem biocatalysis
Co-reporter:Yi Xu, Aitao Li, Xin Jia and Zhi Li
Green Chemistry 2011 vol. 13(Issue 9) pp:2452-2458
Publication Date(Web):18 Jul 2011
DOI:10.1039/C1GC15501F
Novel and efficient one-pot enzymatic and chemo-enzymatic synthetic methods are developed for the asymmetric trans-dihydroxylations of cyclic olefins 1a and 1bvia sequential epoxidation and hydrolysis. The Novozym 435®-mediated epoxidation of cyclohexene 1a and subsequent hydrolysis of the intermediate cyclohexene oxide 2a with resting cells of Sphingomonas sp. HXN-200 in one-pot gave (1R,2R)-cyclohexane diol 3a in 84% ee and 95% conversion. trans-Dihydroxylation of N-benzyloxycarbonyl 3-pyrroline 1b with the same enzymatic system gave the corresponding (3R,4R)-N-benzyloxycarbonyl-3,4-dihydroxy-pyrrolidine 3b in 93% ee and 94% conversion. In the one-pot chemo-enzymatic system, epoxidation of N-benzyloxycarbonyl 3-pyrroline 1b by m-CPBA and subsequent hydrolysis of epoxide intermediate 2b with resting cells of Sphingomonas sp. HXN-200 gave the trans-diol (3R,4R)-3b in 92% ee and 94–97% conversion. While the trans-dihydroxylation of cyclohexene 1a to (1R,2R)-cyclohexane diol 3a is reported for the first time, the trans-dihydroxylation of N-benzyloxycarbonyl 3-pyrroline 1b to (3R,4R)-3b with such an enzymatic or chemo-enzymatic system afforded a much higher product concentration than the same reaction with the system using a microorganism containing the two necessary enzymes. The developed one-pot enzymatic and chemo-enzymatic systems for the asymmetric trans-dihydroxylation of olefins are new, easy to prepare, adjust and operate, are high yielding, complementary to Sharpless asymmetric dihydroxylation and particularly useful for the asymmetric synthesis of cyclic trans-diols.
Co-reporter:Shiyao Dai, Liang Xue, and Zhi Li
ACS Catalysis 2011 Volume 1(Issue 10) pp:1421
Publication Date(Web):September 2, 2011
DOI:10.1021/cs200407n
Novozym 435-catalyzed ring-opening polymerization (ROP) of trimethylene carbonate (TMC) with telechelic hydroxylated poly[(R)-3-hydroxybutyrate] [PHB-diol; Mn = 3000 g/mol (GPC)] as initiator gave di-block poly(HB-co-TMC) with different weight percents of the blocks and Mn of 4400–8700 g/mol (GPC) in 54–89% yield, being the first enzymatic preparation of block poly(ester-co-carbonate). The generality of the novel enzymatic method was demonstrated by the enzymatic ROP of TMC with poly(ε-caprolactone)-diol [PCL-diol; Mn of 4200 g/mol (GPC)] to give A–B–A tri-block-poly(TMC-co-CL-co-TMC) with different weight percents of the blocks and Mn of 7700–10600 g/mol (GPC) in 54–67% yield. The prepared block poly(ester-co-carbonate) with two terminal hydroxyl groups was proven to be a useful starting material for the further preparation of thermoplastic block copolymers. Polymerization of di-block poly(24 wt % HB-co-76 wt % TMC) with methylene diphenyl-4,4′-diisocyanate (MDI) afforded the corresponding polyurethane with Mn of 53 800 g/mol (GPC) in 94% yield. The polymer showed excellent thermoplastic properties (Tm of 144 and 152 °C, Tg of −9 °C, εb of 252%, σmax of 6.37 MPa, and E of 23 MPa), being potentially useful for soft tissue engineering.Keywords: biomaterials; block poly(ester-co-carbonate); enzyme catalysis; poly[(R)-3-hydroxybutyrate]; ring-opening polymerization; trimethylene carbonate;
Co-reporter:Wen Wang, Daniel I. C. Wang and Zhi Li
Chemical Communications 2011 vol. 47(Issue 28) pp:8115-8117
Publication Date(Web):20 Jun 2011
DOI:10.1039/C1CC12685G
Ni-NTA functionalized iron oxide magnetic nanoparticle was synthesized and used to selectively immobilize a his-tagged enzyme from cell free extract as an active and recyclable nanobiocatalyst, where purification and immobilization of the target enzyme were accomplished in one pot.
Co-reporter:Wei Zhang, Weng Lin Tang, Daniel I. C. Wang and Zhi Li
Chemical Communications 2011 vol. 47(Issue 11) pp:3284-3286
Publication Date(Web):01 Feb 2011
DOI:10.1039/C0CC04706F
A novel tandem-biocatalysts system consisting of a monooxygenase-containing microorganism and an alcohol dehydrogenase is developed for the concurrent oxidations of methylene groups to ketones in one pot, providing green, clean and simple access to valuable ketones with high yield, excellent selectivity and efficient cofactor recycling.
Co-reporter:Weng Lin Tang, Zhi Li and Huimin Zhao
Chemical Communications 2010 vol. 46(Issue 30) pp:5461-5463
Publication Date(Web):03 Jun 2010
DOI:10.1039/C0CC00735H
We report the first example of directed evolution of a P450 monooxygenase with inverted enantioselectivity for asymmetric biohydroxylation. The biohydroxylation product of the best mutant 1AF4A has an ee of 83% (R) compared to the wild type's ee of 43% (S).
Co-reporter:Wei Zhang;Weng Lin Tang;Zunsheng Wang
Advanced Synthesis & Catalysis 2010 Volume 352( Issue 18) pp:3380-3390
Publication Date(Web):
DOI:10.1002/adsc.201000266
Abstract
A recombinant Escherichia coli expressing P450pyr monooxygenase of Sphingomonas sp. HXN-200 was developed as a useful biocatalyst for regio- and stereoselective hydroxylations, with no side reaction and easy cell growth. The resting E. coli cells showed an activity of 4.1 U/g cdw and 9.9 U/g cdw for the hydroxylation of N-benzylpyrrolidin-2-one 1 and N-benzyloxycarbonylpyrrolidine 3, respectively, being as active as the wide-type strain. Biohydroxylation of N-benzylpyrrolidin-2-one 1 with the resting cells gave (S)-N-benzyl-4-hydroxypyrrolidin-2-one 2 in >99% ee and 10.8 mM, a 2.6 times increase of product concentration in comparison with the wild-type strain. Biohydroxylation of N-tert-butoxycarbonylpiperidin-2-one 5, N-benzylpiperidine 7 and N-tert-butoxycarbonylazetidine 9 with the E. coli cells afforded the corresponding 4-hydroxypiperidin-2-one 6, 4-hydroxypiperidine 8, and 3-hydroxyazetidine 10 in 14 mM, 17 mM, and 21 mM, respectively. Moreover, hydroxylation of (−)-β-pinene 11 with the recombinant E. coli cells showed excellent regio- and stereoselectivity and gave (1R)-trans-pinocarveol 12 in 82% yield and 4.1 mM, which is over 200 times higher than that obtained with the best biocatalytic system known thus far. The recombinant strain was also able to hydroxylate other types of substrates with unique selectivity: biohydroxylation of norbornane 13 gave exo-norbornaeol 14, with exo/endo selectivity of 95%; tetralin 15 and 6-methoxytetralin 17 were hydroxylated at the non-activated 2-position, for the first time, with regioselectivities of 83–84%.
Co-reporter:Yongzheng Chen Dr.;WengLin Tang;Jie Mou Dr. Dr.
Angewandte Chemie International Edition 2010 Volume 49( Issue 31) pp:5278-5283
Publication Date(Web):
DOI:10.1002/anie.201001772
Co-reporter:Yongzheng Chen Dr.;WengLin Tang;Jie Mou Dr. Dr.
Angewandte Chemie 2010 Volume 122( Issue 31) pp:5406-5411
Publication Date(Web):
DOI:10.1002/ange.201001772
Co-reporter:Shiyao Dai, Jinchuan Wu, Zunsheng Wang, Yongzheng Chen, Zhi Li
Tetrahedron 2010 66(34) pp: 6919-6923
Publication Date(Web):
DOI:10.1016/j.tet.2010.06.039
Co-reporter:Liang Xue, Shiyao Dai, Zhi Li
Biomaterials 2010 31(32) pp: 8132-8140
Publication Date(Web):
DOI:10.1016/j.biomaterials.2010.07.043
Co-reporter:Wen Wang ; Yi Xu ; Daniel I. C. Wang
Journal of the American Chemical Society 2009 Volume 131(Issue 36) pp:12892-12893
Publication Date(Web):August 24, 2009
DOI:10.1021/ja905477j
Magnetic nanoparticles (MNPs) with a core diameter of 30 nm comprising several iron oxide crystals, a poly(glycidyl methacrylate) (PGMA) shell with a thickness of 30 nm, and a surface coated with chloroperoxidase (CPO) were facilely fabricated as a nanobiocatalyst for asymmetric sulfoxidation. The covalently bound CPO did not change the original conformation of the active site and showed the same catalytic activity and enantioselectivity as free CPO for the sulfoxidation of thioanisole to produce (R)-methyl phenyl sulfoxide in >99% ee. The thick PGMA shell significantly increased the stability of the nanobiocatalyst: no loss of the sulfoxidation activity was observed after 11 times of recycling and reuse of the catalyst. Thus, the nanobiocatalyst fabricated here showed the best performance among nanosized biocatalyst particles regarding both the retaining of free enzyme activity and the recycling of catalyst. This is also the first example of a nanobiocatalyst for asymmetric oxidation, and the concept could be generally applicable for fabricating active and recyclable nanobiocatalysts.
Co-reporter:Yi Xu, Na Ranong Bo Jian Khaw and Zhi Li
Green Chemistry 2009 vol. 11(Issue 12) pp:2047-2051
Publication Date(Web):30 Sep 2009
DOI:10.1039/B913077B
A green and efficient oxidation system containing hydrogen peroxide, lactone, and lipase was developed for the epoxidation of alkenes. A variety of alkenes was oxidized with this system, giving 87–95% analytical yield of the corresponding epoxides. The epoxidation occurred vialipase-catalyzed formation of hydroxy peroxy acid from lactone, without release of any harmful short-chain acid and alcohol, and in situ chemical oxidation of alkenes. Both hydrophilic ε-caprolactone and hydrophobic δ-decanolactone were shown to be good substrates to produce hydroxy peracids and good reaction solvents, and the method is suitable for the oxidation in either single phase or two-liquid phase. In comparison with other lipase-mediated oxidation systems, the new oxidation system gave higher yield, higher efficiency, and higher enzyme stability.
Co-reporter:Yi Xu, Xin Jia, Sven Panke and Zhi Li
Chemical Communications 2009 (Issue 12) pp:1481-1483
Publication Date(Web):20 Feb 2009
DOI:10.1039/B820889A
Chiral aryl vicinal diols were obtained in high ee and yield by asymmetric dihydroxylation of arylolefins with tandem biocatalysts: one contains an enantioselective styrene monooxygenase, and the other contains a regioselective epoxide hydrolase.
Co-reporter:Zunsheng Wang;Felicia Lie;Estella Lim;Keyang Li
Advanced Synthesis & Catalysis 2009 Volume 351( Issue 11-12) pp:
Publication Date(Web):
DOI:10.1002/adsc.200900210
Abstract
Cellulosimicrobium cellulans EB-8-4 was discovered by screening of microorganisms as a powerful catalyst for the regio- and stereoselective allylic hydroxylation of D-limonene to (+)-trans-carveol that is a useful and valuable fragrance and flavour compound. Cells of strain EB-8-4 were easily obtained, demonstrated more than 99% regio- and stereoselectivity, showed a specific hydroxylation activity of 4.0 U/g cdw (cell dry weight), and accepted 62 mM D-limonene without inhibition. The hydroxylation was possibly catalyzed by an nicotinamide adenine dinucleotide (NADH)-dependent oxygenase involved in the degradation of aromatic ring during cell growth. 13.4 mM of (+)-trans-carveol were obtained by biohydroxylation of D-limonene with resting cells of C. cellulans EB-8-4, thus being 11 times higher than that obtained with the best biocatalyst known thus far. High conversion and high yield were obtained in the biohydroxylation of 11.6 mM of D-limonene with the resting cells as catalyst in a closed shaking flask, giving 10 mM of (+)-trans-carveol, and 0.30 mM of carvone as the only by-product. Thus, a unique biocatalyst for the regio- and stereoselective allylic hydroxylation of D-limonene and an efficient synthesis of natural identical (+)-trans-carveol by biohydroxylation have been developed.
Co-reporter:Yongzheng Chen;Felicia Lie
Advanced Synthesis & Catalysis 2009 Volume 351( Issue 13) pp:2107-2112
Publication Date(Web):
DOI:10.1002/adsc.200900241
Abstract
Highly enantioselective benzylic hydroxylations of benzene derivatives (1–4) containing reactive functional groups were achieved for the first time with Pseudomonas monteilii TA-5 as biocatalyst, giving the corresponding (R)-benzylic alcohols 5–8 in 93–99% ee as the only products. Preparative biotransformations were demonstrated by the biohydroxylation of 1 and 2 with resting cells of P. monteilii TA-5 to afford (R)-5 in 94% ee and 66% yield and (R)-6 in 94% ee and 56% yield, respectively. The highly enantioselective biohydroxylations represent a simple access to (R)-benzylic alcohols containing reactive functional groups that are useful pharmaceutical intermediates and versatile chiral building blocks.
Co-reporter:Liang Xue, Shiyao Dai and Zhi Li
Macromolecules 2009 Volume 42(Issue 4) pp:964-972
Publication Date(Web):January 23, 2009
DOI:10.1021/ma802437f
Novel biodegradable star poly(ester−urethanes) containing three-arm poly(ε-caprolactone) (PCL) as switching segment were prepared as shape-memory polymers (SMPs) with switching temperature (Ts) around body temperature. PCL-triols with molecular weight (Mn) of 2700−4200 g/mol and Tm of 45−47 °C were synthesized in 55−67% yield by Novozym 435-catalyzed ring-opening polymerization of ε-caprolactone with glycerol as initiator, and their three-arm structures were confirmed by 1H and 13C NMR analysis. Reaction of the PCL-triols with methylene diphenyl 4,4′-diisocyanate isocynate and 1,6-hexanediol gave three-arm PCL-based poly(ester−urethane)s (tPCL-PUs) in 83−92% yield, with 65−75% soft segment. The structure of tPCL-PUs was confirmed by 1H NMR analysis, and the thermal properties were analyzed by DSC with Ts of 36−39 °C. tPCL-PUs showed excellent shape-memory effects at 38 °C during cyclic thermomechanical tensile tests: shape recovery within 10 s, shape fixity rate of 92%, and shape recovery rate of 99%. The novel biodegradable star SMPs are potentially useful in biomedical applications.
Co-reporter:Felicia Lie, Yongzheng Chen, Zunsheng Wang, Zhi Li
Tetrahedron: Asymmetry 2009 Volume 20(Issue 10) pp:1206-1211
Publication Date(Web):5 June 2009
DOI:10.1016/j.tetasy.2009.04.006
A set of 22 toluene- and ethylbenzene-degrading strains were screened for the enantioselective benzylic hydroxylation of indan and tetralin, and Pseudomonas monteilii TA-5 was discovered as an active and selective biocatalyst for such hydroxylations. Cells of P. monteilii TA-5 can be easily grown to a high density and demonstrated a specific hydroxylation activity of 24 U/g cdw (cell dry weight). Conditions for the hydroxylation of indan 1a and tetralin 1b with resting cells of this strain were optimized, to give the corresponding (R)-1-indanol 2a and (R)-1-tetralol 2b in 99% ee and 62–67% yields, respectively. No significant product inhibition was observed, and biohydroxylation with cell-free extracts suggested that the responsible hydroxylase is a soluble enzyme depending on either NADH or NADPH. Preparative biohydroxylation was demonstrated with resting cells as biocatalysts, affording (R)-2a in 99% ee and 65% yield, and (R)-2b in 99% ee and in 63% yield, respectively.(R)-2,3-Dihydro-1H-inden-1-olC9H10O[α]D22=-35.2 (c 1.05, CHCl3)Ee >99%Source of chirality: enzyme catalysisAbsolute configuration: (R)(R)-1,2,3,4-Tetrahydronaphthalen-1-olC9H10O[α]D22=-34.9 (c 1.12, CHCl3)Ee >99%Source of chirality: enzyme catalysisAbsolute configuration: (R)
Co-reporter:Shiyao Dai, Liang Xue, Manfred Zinn and Zhi Li
Biomacromolecules 2009 Volume 10(Issue 12) pp:
Publication Date(Web):November 17, 2009
DOI:10.1021/bm9011634
Enzyme-catalyzed polycondensation for the synthesis of block copolymers was reported for the first time. Thermoplastic block copolyesters containing poly[(R)-3-hydroxybutyrate] (PHB) and poly[(R)-3-hydroxyoctanoate] (PHO) blocks were enzymatically prepared by one- or two-step lipase-catalyzed polycondensation. Novozym 435-catalyzed reaction of PHB-diol (Mn of 3100 g/mol, GPC), PHO-diol (Mn of 3200 g/mol, GPC), and divinyl adipate gave block poly(HB-co-HO) (Mn of 9800 g/mol, GPC) with randomly arranged blocks in 55% yield. In two-step polycondensations, Novozym 435-catalyzed reaction of PHB-diol and divinyl adipate afforded 73% of PHB containing two vinyl ester end groups (Mn of 2700 g/mol, GPC), which was further reacted with PHO-diol in the presence of Novozym 435 to give block poly(HB-co-HO)s (Mn of 8800−14 200 g/mol, GPC) with A−B-type arranged blocks in 55−62% yield. The enzymatically prepared block copolyesters demonstrated Tm of 136−142 °C and 142−153 °C and Tg of −37 to −39 °C and were potentially useful thermoplastic biodegradable and biocompatible materials.
Co-reporter:Xin Jia, Zunsheng Wang, Zhi Li
Tetrahedron: Asymmetry 2008 Volume 19(Issue 4) pp:407-415
Publication Date(Web):4 March 2008
DOI:10.1016/j.tetasy.2007.12.019
The epoxide hydrolase from Sphingomonas sp. HXN-200 catalyzed the enantioselective hydrolysis of racemic 2-, 3-, and 4-chlorostyrene oxides 1–3 to form the corresponding (R)-diols and gave the (S)-epoxides 1–3 in high ee. The reactions were examined with frozen/thawed cells as well as cell-free extracts of Sphingomonas sp. HXN-200 as catalysts in an aqueous, and a two-liquid phase system, respectively. Biotransformation in the two-liquid phase system containing n-hexane as an organic phase showed a higher enantioselectivity than that in the single aqueous phase, due to the reduced non-enzymatic hydrolysis. Hydrolysis of 60 mM 2-chlorostyrene oxide 1 gave 31.3% of (S)-2-chlorostyrene oxide 1 in 98.8% ee with an enantioselectivity factor (E) of 12; hydrolysis of 100 mM 4-chlorostyrene oxide 3 afforded 30.8% of (S)-4-chlorostyrene oxide 3 with 98.6% ee with an E-value of 11. The best results were obtained with the hydrolysis of 3-chlorostyrene oxide 2: biotransformation with 100 mM substrate gave 44.0% of (S)-3-chlorostyrene oxide 2 in 99.0% ee with an E-value of 41; such enantioselectivity is much higher than that of any other known epoxide hydrolases for this reaction; preparative biotransformation demonstrated the efficient synthesis of (S)-3-chlorostyrene oxide 2, an intermediate for the preparation of an IGF-1R kinase inhibitor BMS-536924, with 99.1% ee and 41% isolated yield.(S)-(2-Chlorophenyl)-oxiraneC8H7ClOee = 98.5% (HPLC)[α]D20=+58.2 (c 0.20, CHCl3)Configuration: (S)(S)-(3-Chlorophenyl)-oxiraneC8H7ClOee = 99.1% (HPLC)[α]D20=+11.5 (c 1.61, CHCl3)Configuration: (S)(S)-(4-Chlorophenyl)-oxiraneC8H7ClOee = 99.1% (HPLC)[α]D20=+24.5 (c 1.37, CHCl3)Configuration: (S)
Co-reporter:Shiyao Dai and Zhi Li
Biomacromolecules 2008 Volume 9(Issue 7) pp:
Publication Date(Web):June 10, 2008
DOI:10.1021/bm8001396
Enzymatic modification of a microbial polyester was achieved by the ring-opening polymerization of ϵ-caprolactone (CL) with low-molecular weight telechelic hydroxylated poly[(R)-3-hydroxybutyrate] (PHB-diol) as initiator and Novozym 435 (immobilized Candida antarctica Lipase B) as catalyst in anhydrous 1,4-dioxane or toluene. The ring-opening polymerization was investigated at different conditions with two different types of PHB-diols: PHB-diol(P), containing a primary OH and a secondary OH end groups, and PHB-diol(M), consisting of 91% PHB-diol(P) and 9% PHB-diol containing two secondary OH end groups. The reactions were followed by GPC analyses of the resulting polymers at different time points, and the optimal conditions were established to be 70 °C at a weight ratio of CL/enzyme/solvent of 8:1:24. The ring-opening polymerization of CL with PHB-diol(M) (Mn of 2380, NMR) at the molar ratio of 50:1 under the optimal conditions in 1,4-dioxane gave the corresponding poly[HB(56 wt %)-co-CL(44 wt %)] with Mn (NMR) of 3900 in 66% yield. Polymerization of CL and PHB-diol(P) (Mn of 2010, NMR) at the same condition in toluene gave the corresponding poly[HB(28 wt %)-co-CL(72 wt %)] with Mn (NMR) of 7100 in 86% yield. Both polymers were characterized by 1H and 13C NMR and IR analyses as di-block copolyesters containing a PHB block with a secondary OH end group and a poly(ϵ-caprolactone) (PCL) block with a primary OH end group. NMR analyses and control experiments suggested no formation of random copolymers and no change of the PHB block during the reaction. The enzymatic ring-opening polymerization was selectively initiated by the primary OH group of PHB-diol, whereas the secondary OH group remained as an end group in the final polymers. The thermal properties of the di-block poly(HB-co-CL)s were analyzed by DSC, with excellent Tg values for the elastomer domain: poly[HB(56 wt %)-co-CL(44 wt %)] with Mn (NMR) of 3900 demonstrated a Tg of −57 °C, Tm of 145, 123, and 53 °C; and poly[HB(28wt%)-co-CL(72wt%)] with Mn (NMR) of 7100 gave a Tg of −60 °C, Tm of 147 and 50 °C. Thus, the selective enzymatic ring-opening polymerization with PHB-diol as macro-initiator provides a new method for the preparation of PHB-based block copolymers as biomaterials with good thermoplastic properties and novel structures containing functional end groups.
Co-reporter:Jie Zhang;Bernard Witholt
Advanced Synthesis & Catalysis 2006 Volume 348(Issue 4-5) pp:
Publication Date(Web):2 MAR 2006
DOI:10.1002/adsc.200505439
We have demonstrated, for the first time, the efficient recycling of NADPH in a bioreduction with permeabilized cells of a single microorganism. Permeabilized cells of Bacillus pumilus Phe-C3 containing an NADPH-dependent ketoreductase and a glucose 6-phosphate dehydrogenase (G-6-PDH) were successfully used for the reduction of ethyl 3-oxo-4,4,4-trifluorobutanoate (1) to give (R)-ethyl 3-hydroxy-4,4,4-trifluorobutanoate (2) in 95% ee with the recycling of NADPH for 4220 times from the externally added NADP+. The permeabilized cells were shown to be stable and active for a long period, allowing for high product concentration with high cofactor TTN by continuing the bioreduction with renewed addition of NADP+. This provides with not only a practical synthesis of (R)-2 but also a useful method applicable to many microbial oxidoreductions, since G-6-PDH is a very common enzyme existing in many microorganisms.
Co-reporter:Mojtaba Binazadeh, Iftekhar A. Karimi, Zhi Li
Enzyme and Microbial Technology (7 September 2009) Volume 45(Issue 3) pp:195-202
Publication Date(Web):7 September 2009
DOI:10.1016/j.enzmictec.2009.06.001
Biodegradation of long chain n-alkanes and crude oil with fast rate and high concentration are desirable for bioremediation, especially in heavily oil-polluted areas, and enhanced oil recovery. We discovered Rhodococcus sp. Moj-3449 with such unique abilities by screening microorganisms for the growth on n-hexadecane at 30 mg/mL. The new strain grew very fast on 120 mg/mL of n-hexadecane giving a cell density of 14.7 g cdw/L after only 2 days’ incubation. During the growth with this strain, the oil–water phases were rapidly emulsified, giving rise to tolerance to high alkane concentration (250 mg/mL) and fast growth rate of 0.10–0.20 h−1 for alkane concentration of 1–180 mg/mL. The degraded concentration of n-hexadecane increased linearly with the initial alkane concentration (1–250 mg/mL). Incubation on n-hexadecane at 250 mg/mL for 7 days gave a cell density of 13.5 g cdw/L and degraded 124 mg/mL of n-hexadecane. The strain grew also fast on n-dodecane (C12), n-tetradecane (C14), and n-octadecane (C18), with degradation preference of C14 (=C16) > C12 > C18. Different from many alkane-degrading strains, Rhodococcus sp. Moj-3449 was found to have subterminal oxidation pathway. Rhodococcus sp. Moj-3449 degraded also crude oil fast at 60–250 mg/mL, with a wide range of n-alkanes (C10–C35) as substrates in which C14–C19 are preferred. The degradation ability increased with initial oil concentration from 60 to 150 mg/mL and slightly decreased afterwards. Incubation on 150 mg/mL of crude oil for 7 days degraded 37% of n-alkanes. The outstanding ability of rapidly degrading long chain n-alkanes and crude oil at high concentration makes Rhodococcus sp. Moj-3449 potentially useful for bioremediation and microbial enhanced oil recovery.
Co-reporter:Kaiyuan Tian, Zhi Li
Biochemical Engineering Journal (15 November 2016) Volume 115() pp:30-37
Publication Date(Web):15 November 2016
DOI:10.1016/j.bej.2016.08.002
•Wet cells are used, for the first time, in biodiesel production to give high yield.•Wet cells of E. coli (TLL) can fully convert grease to FAME in 20–30 wt% water.•Direct use of wet cell pellets as catalyst gives 99% FAME yield from waste grease.•Wet cells-process is general, giving high FAME yield from different types of grease.•Wet cells of E. coli (TLL) are recyclable, retaining 55% activity in the 5th cycle.Green and efficient production of biodiesel from cheap and non-edible resources is highly desirable. Here we develop a practical method for the high-yielding one-pot conversion of low-cost waste grease to biodiesel (FAME) by using wet cells of E. coli expressing intracellular Thermomyces lanuginosus lipase (TLL) as catalyst. E. coli (TLL) was genetically engineered and grew easily to a high cell density with the functional expression of TLL. The easily available and easy to handle wet cells were directly used as catalyst for the biotransformation of waste grease to FAME, with water content of 20–30 wt% (based on grease) and stepwise addition of methanol (4:1 molar ratio to grease) as the optimum conditions. Biotransformation of waste grease from Singapore (21 wt% FFA) and Malaysia (9.8 wt% FFA) with 31 wt% wet cells (26 wt% water) gave 99% and 97% FAME yield, respectively. During the biotransformation, the esterification of FFA was faster than the transesterification of triglycerides and the hydrolysis of triglycerides, and nearly no hydrolysis of FAME was observed. Preparative biotransformation was demonstrated to give FAME in >90% yield with a simple isolation procedure. The wet cells were recyclable and retained 55% productivity in the 5th cycle.Download high-res image (150KB)Download full-size image
Co-reporter:Akbar K. Vahidi, Zunsheng Wang, William S. Y. Wong and Zhi Li
Catalysis Science & Technology (2011-Present) 2016 - vol. 6(Issue 16) pp:NaN6293-6293
Publication Date(Web):2016/05/18
DOI:10.1039/C6CY00755D
O-Acetylserine sulfhydrylase (OASS) was immobilized for the first time as an active and recyclable biocatalyst for the conversion of O-acetyl-L-serine (OAS) with a nucleophile to the corresponding useful and valuable β-substituted L-α-amino acid. The simple and efficient immobilization was achieved via affinity attachment, without costly enzyme purification, by direct treatment of cell free extracts (CFE) of a recombinant Escherichia coli strain expressing His-tagged OASS isozyme CysK or CysM with Ni-NTA functionalized iron oxide magnetic nanoparticles. The obtained nanobiocatalysts His-CysK-MNPs (85 nm) and His-CysM-MNPs (85 nm) showed high specific enzyme loading (150 and 94 mg of protein per g of MNPs, respectively) and enzyme loading efficiency (80 and 100%, respectively) and catalyzed the conversion of OAS and pyrazole to β-pyrazol-1-yl-L-alanine (β-PA) with the full activity (2.3 and 1.7 U mg−1 of protein, respectively) of the corresponding free enzyme. His-CysK-MNPs and His-CysM-MNPs were easily separated under magnetic field and demonstrated good recyclability with the retaining of 57% and 54% productivity in the 10th cycle, respectively. Pretreatment of the nanobiocatalysts with 50 mM β-PA released some weakly bound proteins and improved catalyst stability and recyclability. The pretreated His-CysK-MNPs catalyzed the conversion of OAS and pyrazole to produce 1080 mM β-PA with 60% overall conversion over 20 reaction cycles and retained 62% productivity in the 20th cycle. These results indicate the efficient synthesis of the useful and valuable β-PA, and the immobilized OASS could be useful for the synthesis of other β-substituted L-α-amino acids.
Co-reporter:Son Quang Pham, Guillaume Pompidor, Ji Liu, Xiao-Dan Li and Zhi Li
Chemical Communications 2012 - vol. 48(Issue 38) pp:NaN4620-4620
Publication Date(Web):2012/03/05
DOI:10.1039/C2CC30779K
Directed evolution of a monooxygenase to achieve very high enantioselectivity for hydroxylation at non-activated carbon atoms is demonstrated for the first time, where a triple mutant of P450pyr hydroxylase is obtained via determination of enzyme structure, iterative saturation mutagenesis, and high-throughput screening with a MS-based ee assay to increase the product ee from 53% to 98% for the hydroxylation of N-benzyl pyrrolidine to (S)-N-benzyl 3-hydroxypyrrolidine.
Co-reporter:Wen Wang, Daniel I. C. Wang and Zhi Li
Chemical Communications 2011 - vol. 47(Issue 28) pp:NaN8117-8117
Publication Date(Web):2011/06/20
DOI:10.1039/C1CC12685G
Ni-NTA functionalized iron oxide magnetic nanoparticle was synthesized and used to selectively immobilize a his-tagged enzyme from cell free extract as an active and recyclable nanobiocatalyst, where purification and immobilization of the target enzyme were accomplished in one pot.
Co-reporter:Liang Xue, Shiyao Dai and Zhi Li
Journal of Materials Chemistry A 2012 - vol. 22(Issue 15) pp:NaN7411-7411
Publication Date(Web):2012/03/08
DOI:10.1039/C2JM15918J
Elastic shape-memory polymers as self-expendable polymeric drug-eluting stents are developed for the first time, with good mechanical properties, fast self-expansion, and sustained drug release. Novel star block co-polymer PCTOPDs containing hyperbranched poly(ε-caprolactone) (PCL) switching segment and poly(2-oxepane-1,5-dione) (POPD) hard segment were synthesized in high yield and characterized by NMR, GPC, DSC, tensile test and cyclic thermomechanical tensile test. PCTOPD containing 15–27 wt% POPD (PCTOPD-15–27) are found to be non-cytotoxic thermoplastic elastomers (Tm of 39–40 and 120–129 °C, εb of 908–1060%, σm of 12–20 MPa, and E of 60–91 MPa) with good shape-memory properties at 40 °C (Rf of 95–97%, Rr of 97–99%, and shape recovery time of 35 s). The stent made from PCTOPD-27 gives nearly full self-expansion at 37 °C within 45 s. The collapse pressure at a compressive strain of 30% is 1.7 bar. A stent containing 3.0 wt% paclitaxel releases 42% drug linearly in the first 9 days and 67% drug in 30 days with a slower but nearly linear release for the period of 10–30 days. The developed SMP-based drug-eluting stents might be useful in biomedical application such as the treatment of coronary artery disease.
Co-reporter:Weng Lin Tang, Zhi Li and Huimin Zhao
Chemical Communications 2010 - vol. 46(Issue 30) pp:NaN5463-5463
Publication Date(Web):2010/06/03
DOI:10.1039/C0CC00735H
We report the first example of directed evolution of a P450 monooxygenase with inverted enantioselectivity for asymmetric biohydroxylation. The biohydroxylation product of the best mutant 1AF4A has an ee of 83% (R) compared to the wild type's ee of 43% (S).
Co-reporter:Yi Xu, Xin Jia, Sven Panke and Zhi Li
Chemical Communications 2009(Issue 12) pp:
Publication Date(Web):
DOI:10.1039/B820889A
Co-reporter:Thao P. N. Ngo, Wei Zhang, Wen Wang and Zhi Li
Chemical Communications 2012 - vol. 48(Issue 38) pp:NaN4587-4587
Publication Date(Web):2012/03/08
DOI:10.1039/C2CC30953J
Reversible clusters of nanobiocatalysts are developed via non-covalent interaction among enzyme-bound iron oxide magnetic nanoparticles. Dissociation of the clusters by shaking during biotransformation enables high catalytic performance, and re-clustering by stopping shaking after reaction allows for easy magnetic separation. The novel concept is demonstrated with alcohol dehydrogenase RDR for the enantioselective reduction of 7-methoxy-2-tetralone.
Co-reporter:Wei Zhang, Weng Lin Tang, Daniel I. C. Wang and Zhi Li
Chemical Communications 2011 - vol. 47(Issue 11) pp:NaN3286-3286
Publication Date(Web):2011/02/01
DOI:10.1039/C0CC04706F
A novel tandem-biocatalysts system consisting of a monooxygenase-containing microorganism and an alcohol dehydrogenase is developed for the concurrent oxidations of methylene groups to ketones in one pot, providing green, clean and simple access to valuable ketones with high yield, excellent selectivity and efficient cofactor recycling.
Co-reporter:Renliang Huang, Shuke Wu, Aitao Li and Zhi Li
Journal of Materials Chemistry A 2014 - vol. 2(Issue 6) pp:NaN1676-1676
Publication Date(Web):2013/12/02
DOI:10.1039/C3TA14323F
A novel concept integrating supramolecular self-assembly and electrostatic complexation at an aqueous liquid–liquid interface to synthesize stable peptide–polymer hybrid capsules was developed. The concept was further applied for enzyme immobilization to give stable and active biocatalysts with low enzyme leakage and high encapsulation efficiency, enzyme loading, and recyclability.
Co-reporter:Yi Yang, Yu Tse Chi, Hui Hung Toh and Zhi Li
Chemical Communications 2015 - vol. 51(Issue 5) pp:NaN917-917
Publication Date(Web):2014/11/25
DOI:10.1039/C4CC08479A
Directed evolution of P450pyr created I83M/I82T mutant for highly regioselective terminal hydroxylation of n-butanol to 1,4-butanediol, representing the first achievement of this hydroxylation reaction by chemical or enzymatic methods and an unique example of evolving a hydroxylase to switch the substrate acceptance from a hydrophobic to hydrophilic compound.
Co-reporter:Aitao Li, Ji Liu, Son Q. Pham and Zhi Li
Chemical Communications 2013 - vol. 49(Issue 98) pp:NaN11574-11574
Publication Date(Web):2013/10/17
DOI:10.1039/C3CC46675B
A triple mutant of P450pyr monooxygenase (P450pyrTM) catalysed the epoxidation of several para-substituted styrenes as the first enzyme showing high (R)-enantioselectivity and high conversion, demonstrated a broad substrate range, and showed high enantioselectivity for the epoxidation of an unconjugated 1,1-disubstituted alkene, 2-methyl-3-phenyl-1-propene, and a cyclic alkene, N-phenoxycarbonyl-1,2,5,6-tetrahydropyridine, respectively.
Co-reporter:Aitao Li, Shuke Wu, Joseph P. Adams, Radka Snajdrova and Zhi Li
Chemical Communications 2014 - vol. 50(Issue 63) pp:NaN8774-8774
Publication Date(Web):2014/06/12
DOI:10.1039/C4CC03491K
P450tol monooxygenase was discovered as a unique and highly enantioselective enzyme for asymmetric epoxidation of some terminal alkenes containing electron-withdrawing groups and benzylic hydroxylation of several ethylbenzenes giving the corresponding useful and valuable products, such as (R)-2- and 3-substituted styrene oxides, (S)-4-substituted styrene oxides, and (S)-benzylic alcohols, in high ee.
Co-reporter:Ji Liu, Jinchuan Wu and Zhi Li
Chemical Communications 2014 - vol. 50(Issue 68) pp:NaN9732-9732
Publication Date(Web):2014/07/04
DOI:10.1039/C4CC04150J
Enoyl-ACP reductase (FabI) was identified as a non-OYE ‘ene’-reductase for asymmetric reduction of the CC double bond of α, β-unsaturated ketones. Reduction of several 2-alkylidenecyclopentanones with A-FabI and E-FabI gave (R)-2-alkylcyclopentanones in 95–90% and 70–81% ee, respectively. The product ee was improved to 99–98% in high yield by subsequent one-pot biooxidation.
![Oxirane, [3-(trifluoromethyl)phenyl]-, (2S)-](http://img.cochemist.com/ccimg/850500/850402-04-9.png)
![Oxirane, [3-(trifluoromethyl)phenyl]-, (2S)-](http://img.cochemist.com/ccimg/850500/850402-04-9_b.png)
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![Benzene, 1-methoxy-4-[(R)-methylsulfinyl]-](http://img.cochemist.com/ccimg/93400/93381-75-0_b.png)
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![OXIRANE, [3-(TRIFLUOROMETHYL)PHENYL]-, (R)-](http://img.cochemist.com/ccimg/81500/81422-99-3.png)
![OXIRANE, [3-(TRIFLUOROMETHYL)PHENYL]-, (R)-](http://img.cochemist.com/ccimg/81500/81422-99-3_b.png)
![Benzene, [(R)-ethylsulfinyl]-](http://img.cochemist.com/ccimg/51300/51207-25-1.png)
![Benzene, [(R)-ethylsulfinyl]-](http://img.cochemist.com/ccimg/51300/51207-25-1_b.png)
![[(R)-METHYLSULFINYL]BENZENE](http://img.cochemist.com/ccimg/4900/4850-71-9.png)
![[(R)-METHYLSULFINYL]BENZENE](http://img.cochemist.com/ccimg/4900/4850-71-9_b.png)
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![Tungstate(3-),tetracosa-m-oxododecaoxo[m12-[phosphato(3-)-kO:kO:kO:kO':kO':kO':kO'':kO'':kO'':kO''':kO''':kO''']]dodeca-,hydrogen (1:3)](http://img.cochemist.com/ccimg/1400/1343-93-7.png)
![Tungstate(3-),tetracosa-m-oxododecaoxo[m12-[phosphato(3-)-kO:kO:kO:kO':kO':kO':kO'':kO'':kO'':kO''':kO''':kO''']]dodeca-,hydrogen (1:3)](http://img.cochemist.com/ccimg/1400/1343-93-7_b.png)
