Exposing Asymmetry between Monomers in Alzheimer's Amyloid Fibrils via Reductive Alkylation of Lysine Residues

Abstract

Alzheimer's disease amyloid fibrils are composed of the selfassembled 40-43 residue peptide Aβ 1 (Figure 1). Solving the atomic-level structure of these fibrils represents a key step in the study of biochemical processes related to Alzheimer's Disease. Unfortunately, progress toward this goal has been slow because conventional X-ray and NMR structural methods cannot be applied to fibrous protein samples. In developing any structural model of fibrils the space groups and symmetry operators for Aβ monomers are important considerations. In this communication, data is presented which demonstrates that when Aβ is free in solution, the amino terminus and the side chains of both lysine residues (16 and 28) are equally accessible for reductive alkylation. In contrast, when Aβ is self-assembled into fibrils, chemical access to lysines 16 and 28 can be differentiated. Furthermore, fractional alkylation of lysine 28 supports the argument that the "one-dimensional unit-cell" of an amyloid fibril contains at least two non-equivalent Aβ molecules. Together these data offer powerful new constraints for future models of fibril architecture.

Scanning Cysteine Mutagenesis Analysis of Aβ-(1-40) Amyloid Fibrils

“…Whether these consist of subtle microenvironment differences in a largely uniform fibril preparation, or more major differences indicative of multiple conformers of fibrils, is not clear. With respect to other chemical probes, some fibril preparations previously have been shown to exhibit protein environments that behave uniformly in some parts of the structure and variably in other parts (21,22,51).…”

“…Studies by a variety of techniques have provided additional detail about A␤-(1-40) amyloid fibril/protofilament structure and energetics, focusing on the questions of how the A␤ peptide folds to engage that structure and which residues are involved in the H-bonded core. These include EPR of derivatized Cys mutants (20), solid state NMR (18,19), limited proteolysis (21), chemical accessibility of wild type residues (22), hydrogen-deuterium exchange (23)(24)(25)(26)(27)(28), and proline (26), alanine, 3 and disulfide (30) mutagenesis.…”

Alzheimer β-Amyloid Homodimers Facilitate Aβ Fibrillization and the Generation of Conformational Antibodies

“…The relevance of A␤ K28C homodimers to the biophysical behavior of wild-type A␤ is strongly supported by the following published observations. A reductive methylation of Lys-28 slowed the rate of A␤1-40 aggregation but had little effect on protofibril structure (34) and was partially inaccessible to radiomethylation in A␤1-40 fibrils (35). A␤ homodimers play a direct role in A␤ fibril formation: the A␤ fragment 16 -35 was found to form a double-layered hairpin-like structure with the parallel ␤-sheet stabilized by an intrastrand salt bridge (D23-K28) (36,37).…”

The Alzheimer β-amyloid (Aβ1–39) dimer in an implicit solvent

“…Solid-state NMR data also suggest the burial of Lys28 in the fibrils and provide evidence that Lys28 is more protected against alkylation than either Lys16 or the R-amino group in fibrillar A␤ , even though all three amino groups are equally susceptible to alkylation in monomeric A␤ . 40 The atomic positional fluctuations of backbone atoms are displayed in Fig. 6͑a͒ for each residue in A␤ the rest of the peptide but also smaller than observed in the monomer.…”

“…31͒ and the ptraj module of AMBER9. In order to analyze the binding of certain inhibitor peptides with our final dimer configurations, we use the AUTODOCK4.0 program 32 and ClusPro server. 33 The interactions of the target peptides ␤ 1-27 , ␤ [29][30][31][32][33][34][35][36][37][38][39][40][41][42] , and A␤ 1-39 with the inhibitor peptides BSB-KLVFF, 34 LPFFD, 35 LVFFA, 36 and GVVIA 37 ͑here BSB stands for beta sheet breaker, and the other capital letters characterize the amino acid sequence of the peptides in the one-letter-code͒ are calculated and compared to the experimental results available. 34,36,37 The docking runs are performed using the Lamarckian genetic algorithm.…”