Increasing evidence shows that β-amyloid (Aβ) peptides, which are associated with Alzheimer disease (AD), are heavily glycated in patients, suggesting a role of this irreversible nonenzymatic post-translational modification in pathology. Previous reports have shown that glycation increases the toxicity of the Aβ peptides, although little is known about the mechanism. Here, we used the natural metabolic by-product methylglyoxal as a glycating agent and exploited various spectroscopic methods and atomic force microscopy to study how glycation affects the structures of the Aβ40 and Aβ42 peptides, the aggregation pathway, and the morphologies of the resulting aggregates. We found that glycation significantly slows down but does not prevent β-conversion to mature fibers. We propose that the previously reported higher toxicity of the glycated Aβ peptides could be explained by a longer persistence in an oligomeric form, usually believed to be the toxic species.

During their lifecycle, many marine organisms rely on natural adhesives to attach to wet surfaces for movement and self-defense in aqueous tidal environments. Adhesive proteins from mussels are biocompatible and elicit only minimal immune responses in humans. Therefore these proteins have received increased attention for their potential applications in medicine, biomaterials, and biotechnology. The Asian green mussel Perna viridis secretes several byssal plaque proteins, molecules that help anchoring the mussel to surfaces. Among these proteins, protein-5β (Pvfp-5β) initiates interactions with the substrate, displacing interfacial water molecules before binding to the surface. Here, we established the first recombinant expression in Escherichia coli of Pvfp-5β. We characterized recombinant Pvfp-5β, finding that despite displaying a CD spectrum consistent with features of a random coil, the protein is correctly folded as indicated by MS and NMR analyses. Pvfp-5β folds as a β-sheet–rich protein as expected for an epidermal growth factor-like module. We examined the effects of Pvfp-5β on cell viability and adhesion capacity in NIH-3T3 and HeLa cell lines, revealing that Pvfp-5β has no cytotoxic effects at the protein concentrations used and provides good cell-adhesion strength on both glass and plastic plates. Our findings suggest that the adhesive properties of recombinant Pvfp-5β make it an efficient surface-coating material, potentially suitable for biomedical applications including regeneration of damaged tissues.

Amyloid fibril formation is a distinctive hallmark of a number of degenerative diseases. In this process, protein monomers self-assemble to form insoluble structures that are generally referred to as amyloid fibrils. We have induced in vitro amyloid fibril formation of a PDZ domain by combining mechanical agitation and high ionic strength under conditions otherwise close to physiological (pH 7.0, 37 degrees C, no added denaturants). The resulting aggregates enhance the fluorescence of the thioflavin T dye via a sigmoidal kinetic profile. Both infrared spectroscopy and circular dichroism spectroscopy detect the formation of a largely intermolecular beta-sheet structure. Atomic force microscopy shows straight, rod-like fibrils that are similar in appearance and height to mature amyloid-like fibrils. Under these conditions, before aggregation, the protein domain adopts an essentially native-like structure and an even higher conformational stability (DeltaG(U-F)(H2O)). These results show a new method for converting initially folded proteins into amyloid-like aggregates. The methodological approach used here does not require denaturing conditions; rather, it couples agitation with a high ionic strength. Such an approach offers new opportunities to investigate protein aggregation under conditions in which a globular protein is initially folded, and to elucidate the physical forces that promote amyloid fibril formation.