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

Abstract

The glycation-induced functional change of immunoglobulins is of particular interest. The glycation levels of IgG in 10 healthy subjects and 20 diabetic patients with different degrees of metabolic control were studied by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. It reveals the number of glucose molecules that have condensed on the protein, which 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 identification of the most favored glycation sites has been obtained by MALDI analysis of standard and in vitro glycated IgG and plasma protein fraction of a healthy subject after digestion with papain, releasing Fab and Fc fragments of the molecule. Both experiments, as well as molecular modeling of the whole protein, confirm that the most of glucose molecules have condensed on the Fab fragment of IgG, suggesting that the immune deficiency observed in diabetic patients may be explained at the molecular level by a more effective glycation of the Fab fragment, thus inhibiting the process of molecular recognition between antibody and antigen.

Impaired Function of Antibodies to Pneumococcal Surface Protein A but Not to Capsular Polysaccharide in Mexican American Adults with Type 2 Diabetes Mellitus

“…The nonenzymatic glycation of immunoglobulins as a result of hyperglycemia could result in low antigen and receptor binding capacities of these antibodies. It is therefore likely that despite comparable titers, the functionality of the antibody is compromised in diabetics compared to nondiabetics (25,26). Contrary to our findings with PspA, we observed comparable titers of antibodies to all eight capsular polysaccharides tested in individuals with and individuals without diabetes.…”

Advanced Glycation End Products (AGEs) Damaged IgG, a Target for Circulating Autoantibodies in Patients with Type 1 Diabetes Mellitus

“…Glycation of IgG is of special interest due to its influence on the functionality of immunoglobulin's and overall immunocompetence. Several studies showed the extent of glycation of IgG and its influence on the biological functionality of IgG [5][6][7][8][9][10][11][12][13], but the immunological role of AGEs damage IgG (AGE-IgG) in patients with type 1 DM is still not fully understood. Proper extensive clinical investigation of this aspect with a high level of confidence are needed.To meet this need, we hypothesized that AGE-IgG helps to initiate autoimmunity in type 1 diabetes mellitus patients.…”

Demonstration of Glycated Insulin in Human Diabetic Plasma and Decreased Biological Activity Assessed by Euglycemic-Hyperinsulinemic Clamp Technique in Humans

“…It is now becoming increasingly evident that glycation may also compromise the biological activity of functional proteins, including glucose-6-phosphate dehydrogenase (13), aldehyde reductase (14), glutathione reductase (15), Cu-Zn superoxide dismutase (16,17), HDL (18), and both IgM and IgG (19,20). Studies also suggest that glycation of ␤-cell proteins may contribute to defective insulin secretion, and glucokinase has been implicated (21,22).…”