The two major forms of the amyloid-beta (Aβ) peptide found in plaques in patients suffering from Alzheimer's disease, Aβ40 and Aβ42, only differ by two amino acids in the C-terminal region, yet they display markedly different aggregation behavior. The origins of these differences have remained challenging to connect to specific molecular-level processes underlying the aggregation reaction. In this paper we use a general strategy to apply the conventional workflow of chemical kinetics to the aggregation of the Aβ40 peptide to identify the differences between Aβ40 and Aβ42 in terms of the microscopic determinants of the aggregation reaction. Our results reveal that the major source of aggregates in the case of Aβ40 is a fibril-catalyzed nucleation process, the multistep nature of which is evident through its saturation behavior. Moreover, our results show that the significant differences in the observed behavior of the two proteins originate not simply from a uniform increase in all microscopic rates for Aβ42 compared with Aβ40, but rather are due to a shift of more than one order of magnitude in the relative importance of primary nucleation versus fibril-catalyzed secondary nucleation processes. This analysis sheds light on the microscopic determinants of the aggregation behavior of the principal forms of Aβ and outlines a general approach toward achieving an understanding at the molecular level of the aberrant deposition of insoluble peptides in neurodegenerative disorders.protein aggregation | rate law | neurodegeneration | aggregation mechanism

Alzheimer’s disease is an increasingly prevalent neurodegenerative disorder whose pathogenesis has been associated with aggregation of the amyloid-β peptide (Aβ42). Recent studies have revealed that once Aβ42 fibrils are generated, their surfaces strongly catalyse the formation of neurotoxic oligomers. Here we show that a molecular chaperone, a Brichos domain, can specifically inhibit this catalytic cycle and limit Aβ42 toxicity. We demonstrate in vitro that Brichos achieves this inhibition by binding to the surfaces of fibrils, thereby redirecting the aggregation reaction to a pathway that involves minimal formation of toxic oligomeric intermediates. We verify that this mechanism occurs in living brain tissue by means of cytotoxicity and electrophysiology experiments. These results reveal that molecular chaperones can help maintain protein homeostasis by selectively suppressing critical microscopic steps within the complex reaction pathways responsible for the toxic effects of protein misfolding and aggregation.

Nucleation of new peptide and protein aggregates on the surfaces of amyloid fibrils of the same peptide or protein has emerged in the past two decades as a major pathway for both the generation of molecular species responsible for cellular toxicity and for the autocatalytic proliferation of peptide and protein aggregates. A key question in current research is the molecular mechanism and driving forces governing such processes, known as secondary nucleation. In this context, the analogies with other self-assembling systems for which monomer-dependent secondary nucleation has been studied for more than a century provide a valuable source of inspiration. Here, we present a short overview of this background and then review recent results regarding secondary nucleation of amyloid-forming peptides and proteins, focusing in particular on the amyloid β peptide (Aβ) from Alzheimer's disease, with some examples regarding α-synuclein from Parkinson's disease. Monomer-dependent secondary nucleation of Aβ was discovered using a combination of kinetic experiments, global analysis, seeding experiments and selective isotope-enrichment, which pinpoint the monomer as the origin of new aggregates in a fibril-catalyzed reaction. Insights into driving forces are gained from variations of solution conditions, temperature and peptide sequence. Selective inhibition of secondary nucleation is explored as an effective means to limit oligomer production and toxicity. We also review experiments aimed at finding interaction partners of oligomers generated by secondary nucleation in an ongoing aggregation process. At the end of this feature article we bring forward outstanding questions and testable mechanistic hypotheses regarding monomer-dependent secondary nucleation in amyloid formation.

Reaction network starting from monomer mixtures of Aβ40 and Aβ42. Interaction at the level of primary nucleation only accelerates Aβ40 fibril formation. Separate fibrils form as secondary nucleation and elongation are highly specific.

SignificanceThe aggregation of the amyloid-β (Aβ) peptide into amyloid fibrils is associated with Alzheimer’s disease, and several point mutations leading to early-onset disease have been identified in Aβ. By studying the aggregation of five disease-related mutations in vitro, we rationalize their link to familial Alzheimer’s disease. We have determined the effect of mutations on the individual steps of the overall Aβ42 aggregation reaction and find for four of the mutations a significant increase in the rate of self-replication of fibrils, a process that has been linked to the production of toxic oligomeric species. Furthermore, by investigating the nature of the mutation, we determine the importance of the charge and size of specific residues in the aggregation of the wild-type peptide.