Crystal structure of monomeric human β-2-microglobulin reveals clues to its amyloidogenic properties

Abstract

Dissociation of human ␤-2-microglobulin (␤2m) from the heavy chain of the class I HLA complex is a critical first step in the formation of amyloid fibrils from this protein. As a consequence of renal failure, the concentration of circulating monomeric ␤2m increases, ultimately leading to deposition of the protein into amyloid fibrils and development of the disorder, dialysis-related amyloidosis. Here we present the crystal structure of a monomeric form of human ␤2m determined at 1.8-Å resolution that reveals remarkable structural changes relative to the HLA-bound protein.These involve the restructuring of a ␤ bulge that separates two short ␤ strands to form a new six-residue ␤ strand at one edge of this ␤ sandwich protein. Beta-2-microglobulin (␤ 2 m) constitutes the noncovalently bound light chain of the class I human leukocyte antigen (HLA class I). The protein is 99 residues in length and has a seven-stranded ␤ sandwich fold typical of the Ig superfamily (1). In vivo, ␤ 2 m is continuously shed from the surface of cells displaying HLA class 1 molecules into the serum, where it is transported to the kidneys to be degraded and excreted. As a consequence of renal failure, the concentration of ␤ 2 m circulating in the serum increases up to 60-fold (2). Free ␤ 2 m then associates to form amyloid fibrils that typically accumulate in the musculoskeletal system and result in the development of the disorder dialysis-related amyloidosis. The majority of ex vivo ␤ 2 m amyloid is comprised of full-length wild-type ␤ 2 m, although small amounts (Ͻ30%) of modified or truncated forms are found (2-4).As for other amyloidogenic proteins (5-7), mild acidification has been shown to promote formation of amyloid fibrils in vitro from pure ␤ 2 m, both de novo (8-10) and by extension of ex vivo material (11). Under acidic conditions, high yields of amyloidlike fibrils displaying a variety of morphologies form spontaneously in vitro (8-10). Incubation of ␤ 2 m at pH 3.6 results in the formation of a partially unfolded species that assembles into short curly fibrils Ϸ10 nm wide and Ͻ200 nm in length. By contrast, fibrils with a long straight morphology, more reminiscent of ex vivo amyloid, are generated by incubation of aciddenatured forms of the protein at low ionic strengths at pH 2.5 and below, either alone (10) or in the presence of fibrillar seeds (11). Amyloid fibrils of ␤ 2 m have also been generated in vitro at neutral pH by deletion of the N-terminal 6 residues (12); incubation of the full length protein in the presence of Cu 2ϩ ions (13); dialysis of the protein at neutral pH into H 2 O followed by its concentration by drying onto the membrane surface (14); and by the extension of ex vivo material, believed to proceed by the assembly of a rarely populated intermediate onto the ends of existing fibrillar seeds (11, 15). The common theme to emerge from these studies is that conformational rearrangements of the normally highly soluble native protein are required for amyloid fibrils to form.More than 80 crystal structures of h...

Mechanism of amyloidogenesis: nucleation-dependent fibrillation versus double-concerted fibrillation

“…On the contrary, some amyloidogenic proteins retain their native structures before undergoing conformational transition and eventual fibril formation (55). This group of proteins include prion (56), β2-microglobulin (β2m) (57,58), lysozyme (59,60), transthyretin (TTR) (61), and the variable region of immunoglobulin lightchain (VL) (62), which are responsible for mad cow disease (63), dialysis-related amyloidosis (64), hereditary systemic amyloidosis http://bmbreports.org BMB reports (65), senile systemic amyloidosis (61) and light-chain amyloidosis (62), respectively. In addition, apomyoglobin prepared by phase separation using the acid methylethylketone method (66) formed amyloid-like structures upon denaturation in 50 mM sodium borate, pH 9.0, at 65 o C, suggesting that amyloid fibril formation could be considered a generic phenomena of proteins as they experience misfolded structures (67,68).…”

Both the cis - trans equilibrium and isomerization dynamics of a single proline amide modulate β2-microglobulin amyloid assembly

“…1) (12). Several lines of evidence, including detailed folding studies and mutagenesis experiments on β2m, have suggested that cis-to-trans isomerization of Pro32 serves as a trigger to misfolding and subsequent aggregation (6,13).…”

Camelid single-domain antibody fragments: Uses and prospects to investigate protein misfolding and aggregation, and to treat diseases associated with these phenomena

“…b2m is expressed on the surface of nearly all nucleated cells and it adopts an immunoglobulin fold, made up of a seven-stranded b-sandwich linked by a disulphide bridge (Fig. 4E, left part) [88,89]. Although the increased concentration of b2m is a decisive factor in causing DRA, it is not sufficient by itself.…”