Introduction
The quest to determine precise molecular mechanisms by which amyloid beta (Aβ) peptides wreak havoc on the human brain can appear hopeless. These peptides are shapeshifters; they assume countless forms that are often present simultaneously and likely have partially occupied secondary structures (see Deleanu et al.1 and Urban et al.2 for recent reviews). Numerous factors affect Aβ assemblies; e.g., ions and heavy cations, lipids, concentration, time, method of isolation and preparation, location in the body, initial seed conformation, length of the peptide, and oxidation. Also, they appear to interact with up to twenty receptors3, and with several other types of amyloid-forming peptides.
It remains unclear which of their many guises and interactions are the culprits. Much attention has focused on the most visible and stable forms: fibrils within the amyloid plaques that are the hallmark of Alzheimer’s Disease (AD). Their stability has allowed the molecular structure of some forms to be determined experimentally. However, fibrils come in multiple forms: some have U-shaped monomers4-7; others have S-shaped monomers5,8, some have two-fold symmetry along their long axis and others have three-fold symmetry5,8; and most have in-register parallel β-sheets, one has an elongated plus a β-hairpin conformation9, and at least one highly toxic mutant forms antiparallel β-sheets4-7,10. However, evidence is increasing that much smaller assemblies, called oligomers, are more detrimental (reviewed in11-13) and that the longer of the two major forms, Aβ42, is the most toxic12. A subset of the oligomer school contends that Aβ oligomers perturb neuronal signaling and eventually kill neurons by forming transmembrane ion channels13-20.
Many, perhaps most, large Aβ assemblies reflect their origin: i.e., the final structure depends upon the ‘seed’ structure from which it has grown21. Previously we attempted to address these issues by developing atomically explicit models of the structures of Aβ42 hexamers, dodecamers, annular protofibrils, and an ion channel22,23. The starting point for our models was the hypotheses that Aβ42 hexamers can adopt a concentric antiparallel β-barrel structure with a hydrophobic core β-barrel, formed by the last third of the sequence (S3), surrounded by a more hydrophilic β-barrel formed by the first third (S1) and middle third (S2) of the sequence. In these models all monomers have well defined identical conformations and interactions with other monomers.
Several aspects of our hexamer model were unprecedented: (1) six-stranded antiparallel beta-barrels had never been reported. However; Laganowsky et al. 24 have since found that a segment from an amyloid-forming protein, alpha B crystalline, indeed has the six-stranded antiparallel β-barrel motif (which they call Cylindrin), and Do et al. 25 found that several eleven-residue peptides with the sequences of portions of S3 that includes methionine also form this motif. Their calculations confirm our findings that the presence of glycine facilitates packing of aliphatic side chains (especially methionine) in the interior of the barrel. The importance of these residues is supported by findings that mutation of Gly33 to Ala26 or oxidation of Met3527 reduces toxicity and alters oligomerization of Aβ42. (2) Our β structures were antiparallel whereas all known Aβ fibril β-structures were parallel. However, since then an Iowa mutant responsible for some forms of early onset AD has been shown to form fibrils with antiparallel β-sheets10. More important, recent experiments indicate that some Aβ42 oligomers do have an antiparallel β secondary structure that is similar to that of OMPA (an antiparallel β-barrel channel)28-30, and NMR studies of tetramers, octamers31, and 150 kDa oligomers32 indicate that S3 β-strands are antiparallel. Also, antiparallel oligomers are more toxic than those with parallel structures28,30. (3) Concentric β-barrels had never been observed when we proposed the structures. But recent studies have found that the channel-forming toxins Areolysin33 and Lysenin34 do, in fact, contain concentric β-barrels. (4) There was no experimental evidence supporting our proposal that the S1 segments form a β-strand or possibly a β-hairpin. However, subsequently two fibril structures with S-shaped monomers that include S1 have been solved (one with 3-fold symmetry5,8 and one with two-fold symmetry4). In both, the S1 and S2 segments comprise a parallel β-sheet with a bend near the center of S1. We model S1 in two basic conformations: as a continuous β-strand from residues 2-13, and as a β-hairpin with residues A2-H6 forming the first strand (S1a) and residues Y10-H13 forming the second strand (S1b). The β-turn of the hairpin occurs at residues with a high propensity for turn and coil conformations (D7-S8-G9)35, but the exact location and direction of the turn varies among the models. Although often ignored by modelers, numerous findings indicate that alterations within S1 segments affect the toxicity of Aβ and that some mutations within S1 are pathogenic (see 8 and 36 for reviews). Banchelli et al.37 found that Cu++ causes formation of dimers by binding between His6 and His13 or His14 of two Aβ monomers. They concluded that at least portions of adjacent S1 segments are antiparallel (consistent with some of our models) rather than in-register parallel. Also, Tyr10 side chains can cross-link under oxidizing conditions to form dimers38, indicating that they are proximal in some oligomers. (5) There was no experimental evidence that Aβ42 forms β-barrels. However, Serra-Batiste et al. 19 have recently discovered membrane-mimicking conditions under which Aβ42, but not Aβ40 peptides, form a well-defined β-barrel composed of only two monomeric conformations. These results support our proposal that Aβ42 channels contain well-ordered β-barrels resembling those we proposed for oligomers and annular protofibrils.
Experimental constraints used in developing the models presented here were derived primarily from negatively stained electron micrographs of annular protofibrils. Two types of Annular ProtoFibrils have been reported: beaded APFs (bAPFs) that resemble necklaces formed by a string of beads, and smooth APFs (sAPF) that resemble smooth rings39. Portions of electron micrographs used in this study are shown in Fig. 1. These APFs form in the presence of hexane, with bAPFs forming initially from oligomers, then gradually transforming into sAPFs. Also, we have incorporated results of recent solid-state NMR studies of oligomers31,32 in our latest models.