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 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.
Lapolla
A
A
Dipartimento di Scienze Mediche e Chirurgiche, Università di Padova, Italy.
Fedele
D
D
Garbeglio
M
M
Martano
L
L
Tonani
R
R
Seraglia
R
R
Favretto
D
D
Fedrigo
M A
MA
Traldi
P
P
D000368:Aged; D001786:Blood Glucose / Q000378:metabolism; D001798:Blood Proteins / Q000737:chemistry; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism; D005947:Glucose / Q000737:chemistry*; D006801:Humans; D006868:Hydrolysis; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry; D007074:Immunoglobulin G / Q000737:chemistry*; D008958:Models, Molecular; D008970:Molecular Weight; D010206:Papain; D011487:Protein Conformation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D001786:Blood Glucose; D001798:Blood Proteins; D007140:Immunoglobulin Fab Fragments; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D010206:Papain; D005947:Glucose