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Cross-seeding of WT amyloid-β with Arctic but not Italian familial mutants accelerates fibril formation in Alzheimer's disease.

ABSTRACT: Alzheimer's disease (AD) involves the neurotoxic self-assembly of a 40 and 42 residue peptide, Amyloid-β (Aβ). Inherited early-onset AD can be caused by single point mutations within the Aβ sequence, including Arctic (E22G) and Italian (E22K) familial mutants. These mutations are heterozygous, resulting in an equal proportion of the WT and mutant Aβ isoform expression. It is therefore important to understand how these mixtures of Aβ isoforms interact with each other and influence the kinetics and morphology of their assembly into oligomers and fibrils. Using small amounts of nucleating fibril seeds, here, we systematically monitored the kinetics of fibril formation, comparing self-seeding with cross-seeding behavior of a range of isoform mixtures of Aβ42 and Aβ40. We confirm that Aβ40(WT) does not readily cross-seed Aβ42(WT) fibril formation. In contrast, fibril formation of Aβ40(Arctic) is hugely accelerated by Aβ42(WT) fibrils, causing an eight-fold reduction in the lag-time to fibrillization. We propose that cross-seeding between the more abundant Aβ40(Arctic) and Aβ42(WT) may be important for driving early-onset AD and will propagate fibril morphology as indicated by fibril twist periodicity. This kinetic behavior is not emulated by the Italian mutant, where minimal cross-seeding is observed. In addition, we studied the cross-seeding behavior of a C-terminal-amidated Aβ42 analog to probe the coulombic charge interplay between Glu22/Asp23/Lys28 and the C-terminal carboxylate. Overall, these studies highlight the role of cross-seeding between WT and mutant Aβ40/42 isoforms, which can impact the rate and structure of fibril assembly.

SUBMITTER: Liang R

PROVIDER: S-EPMC9243174 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Nanoscale fibrils formed by amyloid peptides have a polymorphic character, adopting several types of molecular structures in similar growth conditions. As shown by experimental (e.g., solid-state NMR) and computational studies, amyloid fibril polymorphism hinders both the structural characterization of Alzheimer's Aβ amyloid protofilaments and fibrils at a molecular level, as well as the possible applications (e.g., development of drugs or biomarkers) that rely on similar, controlled molecular arrangements of the Aβ peptides in amyloid fibril structures. We have explored the use of several contact potentials for the efficient identification of minimal sequence mutations that could enhance the stability of specific fibril structures while simultaneously destabilizing competing topologies, controlling thus the amount of structural polymorphism in a rational way. We found that different types of contact potentials, while having only partial accuracy on their own, lead to similar results regarding ranking the compatibility of wild-type (WT) and mutated amyloid sequences with different fibril morphologies. This approach allows exhaustive screening and assessment of possible mutations and the identification of minimal consensus mutations that could stabilize fibrils with the desired topology at the expense of other topology types, a prediction that is further validated using atomistic molecular dynamics with explicit water molecules. We apply this two-step multiscale (i.e., residue and atomistic-level) approach to predict and validate mutations that could bias either parallel or antiparallel packing in the core Alzheimer's Aβ9-40 amyloid fibril models based on solid-state NMR experiments. Besides shedding new light on the molecular origins of structural polymorphism in WT Aβ fibrils, our study could also lead to efficient tools for assisting future experimental approaches for amyloid fibril determination, and for the development of biomarkers or drugs aimed at interfering with the stability of amyloid fibrils, as well as for the future design of amyloid fibrils with a controlled (e.g., reduced) level of structural polymorphism.

Project description:Neurodegenerative diseases associated with abnormal protein folding and ordered aggregation require an initial trigger which may be infectious, inherited, post-inflammatory or idiopathic. Proteolytic cleavage to generate vulnerable precursors, such as amyloid-beta peptide (Abeta) production via beta and gamma secretases in Alzheimer's Disease (AD), is one such trigger, but the proteolytic removal of these fragments is also aetiologically important. The levels of Abeta in the central nervous system are regulated by several catabolic proteases, including insulysin (IDE) and neprilysin (NEP). The known association of human acetylcholinesterase (hAChE) with pathological aggregates in AD together with its ability to increase Abeta fibrilization prompted us to search for proteolytic triggers that could enhance this process. The hAChE C-terminal domain (T40, AChE(575-614)) is an exposed amphiphilic alpha-helix involved in enzyme oligomerisation, but it also contains a conformational switch region (CSR) with high propensity for conversion to non-native (hidden) beta-strand, a property associated with amyloidogenicity. A synthetic peptide (AChE(586-599)) encompassing the CSR region shares homology with Abeta and forms beta-sheet amyloid fibrils. We investigated the influence of IDE and NEP proteolysis on the formation and degradation of relevant hAChE beta-sheet species. By combining reverse-phase HPLC and mass spectrometry, we established that the enzyme digestion profiles on T40 versus AChE(586-599), or versus Abeta, differed. Moreover, IDE digestion of T40 triggered the conformational switch from alpha- to beta-structures, resulting in surfactant CSR species that self-assembled into amyloid fibril precursors (oligomers). Crucially, these CSR species significantly increased Abeta fibril formation both by seeding the energetically unfavorable formation of amyloid nuclei and by enhancing the rate of amyloid elongation. Hence, these results may offer an explanation for observations that implicate hAChE in the extent of Abeta deposition in the brain. Furthermore, this process of heterologous amyloid seeding by a proteolytic fragment from another protein may represent a previously underestimated pathological trigger, implying that the abundance of the major amyloidogenic species (Abeta in AD, for example) may not be the only important factor in neurodegeneration.

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Cross-seeding of WT amyloid-β with Arctic but not Italian familial mutants accelerates fibril formation in Alzheimer's disease.

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