A Glycosylated Bence Jones Protein and its Autologous Amyloid Light Chain Containing Potentially Amyloidogenic Residues

Abstract

Amyloidosis is characterized by deposition of protein fibrils in various tissues. The wide variety of sequences of both amyloidogenic and non‐amyloidogenic immunoglobulin light chains makes them a unique tool for addressing the importance of primary structure in the formation of insoluble fibrils. In this study, we have determined the primary structure of the κ I immunoglobulin light chain from both the urinary Bence Jones protein and the deposited amyloid fibrils of a patient (MH) with primary amyloidosis. The sequence identity of urinary‐excreted and tissue‐deposited light chains excluded biclonality and somatic mutations and confirmed that the light chain existed in both a soluble and an insoluble form. Several residues have been previously reported to be significantly associated with amyloidogenic κ chains. Many of these were found in the MH sequence, including Asp31, Asn45, Phe49, Gln55, His70, Asn/Gly93 and Pro/Val96, thereby supporting their potential role in fibrillogenesis. In addition, Asn20 and Pro60 of protein MH replaced the normally conserved Thr20 and Ser60. Asn20 was glycosylated in both the Bence Jones and the amyloid fibril protein MH. Cumulative effects of amyloid‐associated residues and glycosylation might have rendered the immunoglobulin light chain MH prone to fibril formation.

Serum amyloid A protein forms a complex with a fragment of Apolipoprotein A-I in the domestic blue fox: a protective mechanism against AA amyloidosis?

“…In positions 6 and 7, both Leu/Val and Gly/Arg, respectively, were seen which in part confirmed the data of tryptic peptides T-1 (pos. [4][5][6][7][8][9][10][11][12][13][14][15][16][17][18] and T-1a (pos. [8][9][10][11][12][13][14][15][16][17][18].…”

“…The purity of the sample was also checked by reverse-phase high performance liquid chromatography (RP-HPLC) as described 16 . Edman degradation was performed before and after treatment of the sample with pyroglutamylaminopeptidase (Boehringer-Manheim and Takara Shuzo Co., Ltd. Japan) according to Mozdzanowski et al 17 .…”

“…Amino acid compositional analysis and Edman degradation were performed as described 16 . The protein was digested with trypsin and protease aureus, and cleaved with cyanogen bromide and BNPS-skatole (2-(2'-Nitrophenylsulfenyl)-3-methyl-3'-bromoindolenine) and the recovered polypeptides purified by reverse-phase HPLC (RP-HPLC) 16,18 . The purity, as well as the quantity of the peptides, was checked by capillary electrophoresis, and amino acid analysis of hydrolysed material.…”

Electrophoretic analysis of Bence Jones proteinuria

“…2-DE and other high resolution electrophoretic methods are also being used increasingly to detect and characterise the carbohydrate constituents of glycoproteins by the use of lectin probes following immunoblotting. This may be of particular interest in studies of nephrotoxicity as BJ proteins are glycosylated [91, 149±152] and differential glycosylation is of pathophysiological significance [153] in dictating tissue uptake of free LC [154] and fibrillogenesis [155].…”

Current and New Perspectives on the Molecular and Cellular Mechanisms of Amyloid Formation and Toxicity in Light Chain Amyloidosis

“…Of the 18 amyloidogenic glycosylated LCs in Stevens study, most of them (13/18) also had other PTMs (including S-cysteinylation, fragmentation, dimerization and S-sulfonation), so a definitive role for glycosylation is difficult to delineate. Other studies also implicated glycosylation as an important characteristic among amyloidogenic proteins (Dwulet et al, 1986;Engvig et al, 1998;Foss et al, 1998;Omtvedt et al, 2000), and AL proteins were found to be glycosylated more frequently than circulating non-amyloidogenic free LCs (Holm et al, 1986;Omtvedt et al, 1997). Despite this evidence, the precise role of this PTM has yet to be determined.…”