Topological investigation of amyloid fibrils obtained from β2‐microglobulin

Abstract

Amyloid fibrils of patients treated with regular hemodialysis essentially consists of β2‐microglobulin (β2‐m) and its truncated species ΔN6β2‐m lacking six residues at the amino terminus. The truncated fragment has a more flexible three‐dimensional structure and constitutes an excellent candidate for the analysis of a protein in the amyloidogenic conformation. The surface topology of synthetic fibrils obtained from intact β2‐m and truncated ΔN6β2‐m was investigated by the limited proteolysis/mass spectrometry approach that appeared particularly suited to gain insights into the structure of β2‐m within the fibrillar polymer. The distribution of prefential proteolytic sites observed in both fibrils revealed that the central region of the protein, which had been easily cleaved in the full‐length globular β2‐m, was fully protected in the fibrillar form. In addition, the amino‐ and carboxy‐terminal regions of β2‐m became exposed to the solvent in the fibrils, whereas they were masked completely in the native protein. These data indicate that β2‐m molecules in the fibrils consist of an unaccessible core comprising residues 20–87 with the strands I and VIII being not constrained in the fibrillar polymer and exposed to the proteases. Moreover, proteolytic cleavages observed in vitro at Lys 6 and Lys 19 reproduce specific cleavages that have to occur in vivo to generate the truncated forms of β2‐m occuring in natural fibrils. On the basis of these data, a possible mechanism for fibril formation from native β2‐m is discussed and an explanation for the occurrence of truncated protein species in natural fibrils is given.