De novo designed peptide-based amyloid fibrils

Abstract

Identification of therapeutic strategies to prevent or cure diseases associated with amyloid fibril deposition in tissue (Alzheimer's disease, spongiform encephalopathies, etc.) requires a rational understanding of the driving forces involved in the formation of these organized assemblies rich in ␤-sheet structure. To this end, we used a computer-designed algorithm to search for hexapeptide sequences with a high propensity to form homopolymeric ␤-sheets. Sequences predicted to be highly favorable on this basis were found experimentally to self-associate efficiently into ␤-sheets, whereas point mutations predicted to be unfavorable for this structure inhibited polymerization. However, the property to form polymeric ␤-sheets is not a sufficient requirement for fibril formation because, under the conditions used here, preformed ␤-sheets from these peptides with charged residues form well defined fibrils only if the total net charge of the molecule is ؎1. This finding illustrates the delicate balance of interactions involved in the formation of fibrils relative to more disordered aggregates. The present results, in conjunction with x-ray fiber diffraction, electron microscopy, and Fourier transform infrared measurements, have allowed us to propose a detailed structural model of the fibrils. T he ability of soluble proteins or protein fragments to convert spontaneously into amyloid fibrils under some circumstances is a very important biological phenomenon, because this process is related to a range of human disorders such as spongiform encephalopathies, Alzheimer's disease, etc. (1, 2). Although soluble precursors of amyloidogenic proteins do not have any obvious sequence homology or common folding patterns, x-ray fiber diffraction data indicate that all amyloid fibrils share a characteristic cross-␤-structure (3, 4). This finding suggests that the key elements of the fibril formation process may be common to all proteins and that, therefore, a highly simplified system that is able to polymerize into ␤-sheets can offer additional insights into the molecular details of amyloid fibril formation. Up till now, the search for such simple model systems has been based mainly on empirical approaches such as, for example, the identification of fragments of amyloidogenic proteins that can self-assemble into amyloid fibrils (5-8). Some rational designs based on alternating hydrophobic and hydrophilic residues have been found to have some success in identifying the interactions behind ␤-sheet polymerization (9). Although such systems have helped to clarify many aspects of this process, our detailed understanding of the factors promoting amyloid formation remains limited.To probe the interactions driving ␤-sheet aggregation, we have investigated the effect of specific residues on the propensity of a given sequence to form amyloid fibrils. To this end, we have used a computer-based method (10-12) to design a series of self-associating hexapeptides able to form polymeric ␤-sheet structures arranged in a similar fashion to that assu...

Kinetic control of dimer structure formation in amyloid fibrillogenesis

“…The system was enclosed by a reflecting sphere of diameter 90 Å (this corresponds to a concentration of 8.7 mM, which is only an order of magnitude larger than the highest concentration used in ref. 26) (Fig. 1a; see also Supporting Text).…”

“…The other peptides are designed sequences used in the study of fibril formation and peptide self-assembly. A␤ 7 , K I , and KFE8 form antiparallel ␤-sheet structures (26)(27)(28), whereas K L forms ␤-sheet aggregates but not fibrils (26). Several peptides were studied to determine sequence-independent characteristics of the dimerization process, because, as already mentioned, many different proteins and peptides can form amyloid fibrils (23,29).…”

“…All peptides were acetylated and amidated at N and C termini. For K I and K L , the Glu residue was protonated to accommodate for low pH condition at which the experiment was performed (26). To generate a dimerization trajectory, we first ran a singlepeptide simulation at 298 K for 1 ns to obtain an average monomer structure.…”

Hydrophobic Cooperativity as a Mechanism for Amyloid Nucleation

“…By examining seven other similar sequences Serrano et al were able to show that peptides with a net charge of ±1 had a marked preference for fibril formation over their electrically neutral counterparts. 18 We performed simulations on the STVIYE system with both protonated and deprotonated glutamate to explore the molecular basis for this trend. Simulations were performed using replica exchange molecular dynamics (REMD) with the all-atom CHARMM force field 19 and a generalized Born (GB) implicit solvent model.…”

Monitoring disappearance of monomers and generation of resistance to proteolysis during the formation of the activation domain of human procarboxypeptidase A2 (ADA2h) amyloid fibrils by matrix-assisted laser-desorption ionization–time-of-flight-MS

“…Taken together, these results suggest that, probably, α-helix 2 plays an important role in the formation of the amyloid fibril. In this respect, it is interesting to point out that the recent study by Dobson and coworkers [22] has shown that only two small regions in a large protein affect the formation of amyloid fibrils and that designed short hexapeptides can form amyloid fibrils, which cannot be distinguished from the ones made by large proteins [25].…”