There is compelling evidence that small oligomeric aggregates, emerging during the assembly of amyloid fibrils and plaques, are important molecular pathogens in many amyloid diseases. While significant progress has been made in revealing the mechanisms underlying fibril growth, understanding how amyloid oligomers fit into the fibril assembly process, and how they contribute to the pathogenesis of amyloid diseases, has remained elusive. Commonly, amyloid oligomers are considered to be metastable, early-stage precursors to fibril formation that are either on- or off-pathway from fibril growth. In addition, amyloid oligomers have been reported to colocalize with late-stage fibrils and plaques. Whether these early and late-stage oligomer species are identical or distinct, and whether both are relevant to pathogenesis remains unclear. Here we report on the formation of two distinct oligomer species of lysozyme, formed either during the early or late-stages of in vitro fibril growth. We further observe that the pH change from in vitro growth conditions to cell media used for toxicity studies induced distinct mesoscopic precipitates, two of which resemble either diffuse or neuritic plaques seen in Alzheimer's histology. Our biophysical characterization indicates that both oligomer species share morphological and tinctorial features considered characteristic for amyloid oligomers. At the same time, their sizes, morphologies, their immunostaining, detailed tinctorial profiles and, most prominently, their biological activity are clearly distinct from each other. Probing the conditions promoting the formation of these two distinct oligomer species suggests distinct roles of charge interactions, hydrophobicity and monomer flexibility in directing oligomer assembly.
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