ANALYSES OF IGA1 HINGE GLYCOPEPTIDES IN IGA NEPHROPATHY BY MATRIX-ASSISTED-LASER-DESORPTION IONIZATION TIME-OF-FLIGHT MASS-SPECTROMETRY/

This study was performed to analyze the structural variety of O-glycan s on the IgA1 hinge in IgA nephropathy (IgAN). The IgA1 fragments cont aining the hinge glycopeptide (33-mer hinge peptide core (HP) + O-glyc ans) were separated from 13 IgAN patients, eight healthy control subje cts, and 11 patients with other primary glomerulonephritides by pyridy l-ethylation, trypsin treatment, and Jacalin affinity chromatography. Because of the use of Jacalin, only the Gal beta 1-3GalNAc residue con taining IgA was analyzed, The molecular weights (MW) of the IgA1 fragm ents treated by the following sequential treatment by exoglycosidases were estimated using matrix-assisted laser desorption/ionization time- of-flight mass spectrometry: (1) Sialidase treatment: the MW of the tw o observed peaks A and B were compatible with (A) HP + 4GalNAc + 4Gal and (B) HP + 5GalNAc + 4Gal, (2) Sialidase and galactosidase: the MW o f the two identified peaks a and b were consistent with (a) HP + 4GalN Ac and (b) HP + 5GalNAc, (3) Sialidase, galactosidase, and alpha-N-ace tylgalactosaminidase. All subjects revealed one peak, indicating the 3 3-mer IgA1 hinge peptide core. The intensity rate of peak B/A was sign ificantly decreased in the IgAN group (mean +/- SD, 1.01 +/- 0.08) com pared with the negative control subjects (healthy group, 1.15 +/- 0.06 , P = 0.0048; other glomerulonephritis group, 1.13 +/- 0.10, P = 0.004 9; Scheffe's F test). These results suggested the presence of a defect in the Gal and/or GalNAc residues in the IgA1 hinge, glycopeptides in IgAN.