Fluorometric and mass spectrometric analysis of nonenzymatic glycosylated albumin

Albumin is the major transport protein in blood and intramolecular movement contributes to this function. Nonenzymatic glycosylation (NEG) of albumin occurs in diabetes and, in this study, fluorometric methods were used to de termine the effect of increasing levels of NEG upon intramolecular movement in human serum albumin. Low levels of NEG significantly reduced and left-s hifted Trp fluorescence, reduced quenching by acrylamide and inhibited pene tration of bis-ANS, while these changes became only modestly more pronounce d at higher levels of NEG. Mass spectrometry of tryptic and CNBr NEG-HSA fr agments identified potential glycosylation sites and demonstrated only late glycosylation of the C- and N-terminal regions of the protein. Similar cha nges in diabetes may contribute to altered transport function in these pati ents, (C) 2001 Academic Press.