Infectious mammalian prions can be formed de novo from purified recombinant prion protein (PrP) substrate through a pathway that requires the sequential addition of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) and RNA cofactor molecules. Recent studies show that the initial interaction between PrP and POPG causes widespread and persistent conformational changes to form an insoluble intermediate species, termed PrPInt1. Here, we characterize the mechanism and functional consequences of the interaction between POPG and PrP. Negative-stain electron microscopy of PrPInt1 revealed the presence of amorphous aggregates. Pull-down and photoaffinity label experiments indicate that POPG induces the formation of a PrPC polybasic-domain-binding neoepitope within PrPInt1. The ongoing presence of POPG is not required to maintain PrPInt1 structure, as indicated by the absence of stoichiometric levels of POPG in solid-state NMR measurements of PrPInt1. Together, these results show that a transient interaction with POPG cofactor unmasks a PrPC binding site, leading to PrPInt1 aggregation.

A surprisingly strong spin rate dependence of 15N and 13C NMR T1 times in magic angle spinning experiments on solid peptides is demonstrated. Using a variety of isotopomers, the phenomenon is shown to be the result of chemical shift anisotropy-mediated spin diffusion. This effect has the potential to be used to detect long-range distance constraints in macromolecular systems.