| Abstract
| - β2-microglobulin (β2m) forms amyloid fibrils that deposit in the musculo-skeletal system inpatients undergoing long-term hemodialysis. How β2m self-assembles in vivo is not understood, sincethe monomeric wild-type protein is incapable of forming fibrils in isolation in vitro at neutral pH, whileelongation of fibril-seeds made from recombinant protein has only been achieved at low pH or at neutralpH in the presence of detergents or cosolvents. Here we describe a systematic study of the effect of 11physiologically relevant factors on β2m fibrillogenesis at pH 7.0 without denaturants. By comparing theresults obtained for the wild-type protein with those of two variants (ΔN6 and V37A), the role of proteinstability in fibrillogenesis is explored. We show that ΔN6 forms low yields of amyloid-like fibrils at pH7.0 in the absence of seeds, suggesting that this species could initiate fibrillogenesis in vivo. By contrast,high yields of amyloid-like fibrils are observed for all proteins when assembly is seeded with fibril-seedsformed from recombinant protein at pH 2.5 stabilized by the addition of heparin, serum amyloid Pcomponent (SAP), apolipoprotein E (apoE), uremic serum, or synovial fluid. The results suggest that theconditions within the synovium facilitate fibrillogenesis of β2m and show that different physiologicalfactors may act synergistically to promote fibril formation. By comparing the behavior of wild-type β2mwith that of ΔN6 and V37A, we show that the physiologically relevant factors enhance fibrillogenesis bystabilizing fibril-seeds, thereby allowing fibril extension by rare assembly competent species formed bylocal unfolding of native monomers.
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