Glycation of human beta 2-microglobulin in patients with hemodialysis-associated amyloidosis: identification of the glycated sites.
beta 2-Microglobulin (beta 2M) is a major component forming amyloid deposits in patients with hemodialysis-associated amyloidosis (HAA), a serious complication of long-term hemodialysis. Recently, we demonstrated that beta 2M modified with the Maillard reaction is a definite constituent of amyloid deposits in patients with HAA. Our further study demonstrated that this modified beta 2M induces not only chemotaxis of monocytes but also secretion of tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6 from macrophages, suggesting the potential link of glycation of beta 2M by the Maillard reaction to the pathogenesis of HAA. The present study was undertaken to identify the glycated site(s) of beta 2M purified from long-term hemodialysis patients as well as beta 2M incubated with glucose in vitro. Borotritide-treated beta 2M was cleaved by endoproteinase Lys-C, and peptides were isolated by reverse-phase high-performance liquid chromatography, followed by amino acid sequence analysis and fast atom bombardment mass spectrometry to identify the glycated site. The glycated sites of beta 2M formed in vivo were found to be almost the same as those of glycated beta 2M in vitro. The primary glycated site was the alpha-amino group of the amino terminal isoleucine. Other minor sites were the epsilon-amino groups of Lys-19, -41, -48, -58, -91, and -94. Computer graphics of the three-dimensional structure of beta 2M suggested that the high specificity for the glycated site at Ile-1 may be explained by its high solvent accessibility and the nearby imidazole group of His-31 as an acid-base catalyst of the Amadori rearrangement.
Miyata
T
T
Department of Internal Medicine, Branch Hospital, Nagoya University School of Medicine, Japan.
Inagi
R
R
Wada
Y
Y
Ueda
Y
Y
Iida
Y
Y
Takahashi
M
M
Taniguchi
N
N
Maeda
K
K
D000328:Adult; D000595:Amino Acid Sequence; D000686:Amyloidosis / Q000209:etiology / Q000378:metabolism*; D001798:Blood Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001847:Bone Diseases / Q000209:etiology / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D005260:Female; D006023:Glycoproteins; D006031:Glycosylation; D006801:Humans; D066298:In Vitro Techniques; D007375:Interleukin-1 / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D007592:Joint Diseases / Q000209:etiology / Q000378:metabolism*; D008264:Macrophages / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D006435:Renal Dialysis / Q000009:adverse effects*; D015183:Restriction Mapping; D016339:Spectrometry, Mass, Fast Atom Bombardment; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D001613:beta 2-Microglobulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000093422:Glycated Serum Proteins
D001798:Blood Proteins; D006023:Glycoproteins; D007375:Interleukin-1; D015850:Interleukin-6; D014409:Tumor Necrosis Factor-alpha; D001613:beta 2-Microglobulin; D000093422:Glycated Serum Proteins