抄録
We studied the effects of noneyzymatic glycation on human antithrombin III (AT-Ill). In vitro, purified human AT-III (15U/ml) was incubated with glucose (0, 100, 400, 1, 000mg/dl) at 37°C for 2 weeks. AT-III was significantly glycated and then progressive activity and heparin cofactor activity of glycated AT-III decreased, while the levels of AT-III antigen had not changed. When glycated AT-III (1U/ml) was incubated with thrombin (250NIHU/ml) for various times, the amount of complex formed was less than control (control: 4.3-10.6mg/ml, glycated AT-III: 0.4-6.2mg/ml, p<0.01). In the presence of heparin, glycated AT-III (treated with 1g/dl glucose) was only formed a complex of high molecular weight (M.W.) from SDS-PAGE. The amount of complex formed was the same degree both control and glycated AT-III (treated with 0-400mg/dl glucose). But in the glycated AT-III (treated with 1g/dl glucose), the pattern of complex formation was different from control because of measurable complexes of M.W from 78, 000 to 81, 500 were detected in this method.
On crossed immunoelectrophoresis containing heparin in the first dimension agarose, control AT-III demonstrated AT-III as 2 peaks including the fast moving main peak. On the contrary, glycated AT-III showed 2 main peaks including the slow moving peak, suggesting that the glycated AT-III has little affinity for heparin. We attached a guanidine group to the lysyl residues of AT-III by treating it with O-methylisourea. The modified AT-III was not glycated and then progressive activity had not changed and heparin cofactor activity decreased.
From these observations, it is suggested that lysyl residues of AT-III are essential in nonenzymatic glycation and nonenzymatic glycation and nonenzymatic glycation causes a conformational change which results in a poor formation of complex, decreasing AT-III activity.
