INVESTIGATIONS ON PROTEIN NONENZYMATIC GLYCATION BY A NEW AND EFFECTIVE-MASS SPECTROMETRIC METHOD

Protein nonenzymatic glyco-oxidation has been studied either in vitro or ill who by matrix assisted laser desorption/ionization (MALDI). Bov ine serum albumin and ribonuclease have been incubated In pseudophysio logical conditions with different sugar (glucose or fructose) concentr ations. Mass increases of incubated proteins have been determined by M ALDI and related to different numbers of sugar molecules condensed on the active sites of the protein. The kinetics of glyco-oxidation has b een determined for the various sugar concentrations showing in all cas es the reaching of a steady-state. corresponding to the complete satur ation of the reactive sites. The same methodology has been applied in the study of human serum albumin from healthy subjects and weil-contro lled and badly controlled diabetic patients. Also in this case, clear differences have been evidenced among the three groups of subjects und er investigation. In the case of badly controlled patients, the molecu lar weight of human serum albumin shows increases with respect to genu ine human serum albumin, in the range 439-2403, corresponding to the c ondensation on that protein from 3 to 15 glucose units. (C) 1996 Acade mic Press, Inc.