In vitro glycation of human immunoglobulin G.
BACKGROUND
Glucose can covalently bind to the proteins by nonenzymatic process often termed glycation. Glycation of IgG is of special interest due to its possible influence on the functionality of immunoglobulins and overall immunocompetence.


METHODS
The glycation of IgG was studied using radioactive D-[U-14C]-glucose.


RESULTS
The kinetics, concentration/temperature dependence and distribution of glycation binding sites of human IgG were studied under various conditions. (a) In average, 0.7 glucose molecules were found to be bound per IgG after 90 days of incubation at 37 degrees C with normal serum IgG and glucose concentrations, and 3.1 molecules after incubation with glucose in the concentration characteristic for diabetics. (b) Incubation of the same solutions for 270 days at 4-8 degrees C resulted in binding of 0.3 and 0.8 glucose molecules per IgG, respectively. (c) After 90 days at 37 degrees C in the presence of 0.56 mol/l of glucose, IgG was glycated with averagely 47 glucose molecules per IgG and with 9 after 270 days at 4-8 degrees C. (d) At very high glucose concentration (1.67 mol/l) in concentrated IgG solution (87 g/l), the molar ratio glucose/IgG reached 41 after incubation at 60 degrees C for 10 h.


CONCLUSIONS
Glycation of IgG with glucose nearly resembled the first order reaction kinetics. No preferential glycation sites were found as bound glucose was equally distributed throughout F(ab)2 and Fc fragments as well as on the light and heavy immunoglobulin chains.
Vrdoljak
Anto
A
Institute of Immunology, Rockefellerova 2, HR-10 000 Zagreb, Croatia. avrdoljak@imz.hr
Trescec
Anda
A
Benko
Bojan
B
Hecimovic
Dragica
D
Simic
Mirjana
M
D004355:Drug Stability; D005947:Glucose / Q000737:chemistry; D006801:Humans; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G / Q000737:chemistry*; D016756:Immunoglobulins, Intravenous / Q000737:chemistry; D007202:Indicators and Reagents; D019999:Pharmaceutical Solutions; D013242:Sterilization; D013696:Temperature
D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D016756:Immunoglobulins, Intravenous; D007202:Indicators and Reagents; D019999:Pharmaceutical Solutions; D005947:Glucose