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Stacked sets of parallel, in register beta-strands of beta-2-microglobulin in amyloid fibrils revealed by site-directed spin labelling and chemical labelling

By C. L. Ladner, M. Chen, David Smith, G. W. Platt, S. E. Radford and R. Langen

Abstract

β2-microglobulin (β2m) is a 99-residue protein with an immunoglobulin fold that forms β-sheet rich amyloid fibrils in dialysisrelated amyloidosis. Here the environment and accessibility of side-chains within amyloid fibrils formed in vitro from β2m with a longstraight morphology are probed by sitedirected spin labelling and accessibility to modification with N-ethyl maleimide using 19 site-specific cysteine variants. Continuous wave electron paramagnetic resonance (CW-EPR) spectroscopy of these fibrils reveals a core predominantly organized in a parallel, inregister arrangement, by contrast with other β2m aggregates A continuous array of parallel, in-register β-strands involving most of the polypeptide sequence is inconsistent with the cryo-electron microscopy (cryo-EM) structure which reveals an architecture based on subunit repeats. To reconcile these data, the number of spins in close proximity required to give rise to spin exchange was determined. Systematic studies of a model protein system indicated that juxtaposition of four spin labels is sufficient to generate exchange narrowing. Combined with information about side-chain mobility and accessibility we propose that the amyloid fibrils of β2m consist of ca. six β2m monomers organized in stacks with a parallel, in-register array. The results suggest an organization more complex than the accordion-like β- sandwich structure commonly proposed for amyloid fibrils

Publisher: American Society for Biochemistry and Molecular Biology
Year: 2010
OAI identifier: oai:shura.shu.ac.uk:1870
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