Matrix-assisted laser desorption/ionization mass spectrometry, enzymatic digestion, and molecular modeling in the study of nonenzymatic glycation of IgG

The glycation-induced functional change of immunoglobulins is of particular interest. The glycation levels of IgG in 10 healthy subjects and 20 diabet ic patients with different degrees of metabolic control were studied by mat rix-assisted laser desorption/ionization (MAT,DI) mass spectrometry. It rev eals the number of glucose molecules that have condensed on the protein, wh ich range from 1 to 5 for healthy subjects, from 5 to 9 for well controlled diabetic patients, and from 10 to 25 for poorly controlled ones. The ident ification of the most favored glycation sites has been obtained by MALDI an alysis of standard and in vitro glycated IgG and plasma protein fraction of a healthy subject after digestion with papain, releasing Fab and Fc fragme nts of the molecule. Both experiments, as well as molecular modeling of the whole protein, confirm that the most of glucose molecules have condensed o n the Fab fragment of IgG, suggesting that the immune deficiency observed i n diabetic patients may be explained at the molecular level by a more effec tive glycation of the Fab fragment, thus inhibiting the process of molecula r recognition between antibody and antigen. (C) 2000 American Society for M ass Spectrometry.