2.3 Random coil Oligomers with alpha-helical content
The AlphaFold2 machine-learning approach is based on protein data bank (PDB) templates, sequence alignments, co-evolution rules and multiple algorithms to design a protein-specific potential of mean force. AlphaFold2 success stories include the prediction of single domain protein structures,55 and most transmembrane protein structures.56 AlphaFold2 limitations to predict very accurately the structures of protein – protein (peptide) complexes57,58 and generate conformational heterogeneity59 were reported.
At the date of the present study, the PDB contained about 200,000 structures.60 The most striking AlphaFold2 result for the structures of Aβ42 dimers up to hexamers is the prediction of α-helix topologies for all species in addition to β-rich topologies.54 The AlphaFold2 structures are shown in Figure 1. While the dimer displays an antiparallel helix bundle spanning the C-terminus (Figure 1A), all higher aggregates display parallel helix bundles spanning the C-terminal residues 29-39. (Figures 1B-E). These α-rich oligomers are supported indirectly by numerous experiments on Aβ and many other amyloid polypeptides.
CD experiments on Aβ42 and Aβ40 peptides in pure buffer give 19% and 32% of α-helix structure after 4 days of incubation.61 Addition of trifluoroethanol suggested α-helical intermediates during Aβ assembly,62 and addition of low solvent polarity stabilized partial α-helical structures and accelerated Aβ40 amyloid fibrillation.63Pyroglutamate-modified pEAβ(3-42) aggregation also pointed to α-helical intermediates, stabilized by parallel C-terminus interactions, each monomer forming a helix-turn-helix spanning residues 10-23 and 30-36.64
Slow nucleation of short polyglutamine-containing Huntingtin fragments via α-helix-rich oligomers and inhibition of amyloid structure in a Huntingtin fragment by targeting α-helix-rich oligomers were also reported experimentally.65,66 Using computational and experimental approaches, human islet amyloid polypeptide (hIAPP) fragment 8-20 fibril formation starts from isolated helical monomers, helical dimers to hexamers, followed by the conversion to β at the hexamer level.67 PolyQ-Aβ30-42 peptides at µM concentration suggested an aggregation triggered by a rapid formation of α-helical oligomers mediated by the C-terminal residues, as assessed by CD and FTIR (Fourier Transformed Infrared) spectroscopies.68 Infrared nanospectrometry monitored a α-to-β transition during the self-assembly of the N-terminal Josephin domain of ataxin 3.69 The conversion of rationally designed α-helical peptides to amyloid fibrils and the oligomerization of natural hexapeptides into amyloid fibrils through α-helical oligomers are also well established.70,71 Overall, there are many experiments reporting a minor population of partially folded helical oligomers during amyloid fibril formation.72,73
Additionally, a rational design of α-helical peptide inhibitors targeting Aβ40 surface reduces the generation of toxic Aβ toxic oligomers.74 Helical peptide foldamers and peptidometics were found dual inhibitors of Aβ and hIAPP fibrillization.75 Alpha-helix mimetics, which induce α-helicity in Aβ using NMR and CD, inhibit the seed-catalyzed aggregation of Aβ.76 Based on ion mobility spectrometry – mass spectrometry combined to MD simulations, it was suggested that Aβ C-terminal interactions play a key role in their inhibitory activity.77 Finally, it was found that Aβ25-35 peptide forms early stage helical conformations by CD and Raman spectroscopic techniques, and carvedilol inhibits Aβ25-35 fibrillation.78
Computationally, AlphaFold2 α-helical tetramer and hexamer structures are very stable using CHARMM36m-TIP3P modified and AMBER99SB-DISP for 0.3 µs MD simulations at 310 K.54 Transient formation of helical conformations differing from helix bundles was reported by numerous simulations of Aβ40 and Aβ42 oligomers,7,28,46,79 but a recent simulation proposed that conformations with α-helical structure have a high propensity to initiate Aβ42 aggregation.80 Finally, it should be noted that the helix propensity of amyloid peptides is a fundamental requirement to fulfill the lipid-chaperon model,81 and helical intermediates during amyloid formation are catalysed by membranes.36,72