Anti-thrombine III

In hormone replacement therapy, the risk of deep vein thrombosis is increased by a factor of 2-4 (35-37). The absolute increase in the treated population as a whole is low, with about one case of venous thromboembolism in 5000 women-years of use of hormone replacement therapy. However, in the subgroup with pre-existing risk factors, such as obesity, varicose veins, smoking, and a prior history of venous thromboembolism or superficial thrombophlebitis, the increase in risk from hormone replacement therapy can be substantial among these women are those with a genetic predisposition to thrombosis, generally due to some form of thrombophilia, such as deficiency of the coagulation inhibitors protein S, protein C, or anti thrombin III. In any of these subjects thrombosis can occur early in hormone replacement therapy. However, this tendency to early occurrence of deep vein thrombosis also seems to be present in all those who take hormone replacement therapy. 

Heparin acts by binding to anti thrombin III, which serves as a major inhibitor of serine protease clotting enzymes. Abruptly ending heparin treatment can be hazardous because of reduced levels of antithrombin III. Coumarins, typified by warfarin, are structurally similar to vitamin K, which plays an important role in blood coagulation. By interfering with the function of vitamin K, vitamin K-dependent proteins such as clotting factors VII, IX, X and prothrombin are reduced. 

Thrombin and factor Xa that have escaped into the blood flow are both inhibited by serpins, anti-thrombin III (ATIII) and heparin cofactor II (HCII). These proteins bind to heparin sulfate or dermatan sulfate (Sect. 6.3.1), glycosaminoglycans which are secreted onto the luminal surface of healthy endothelial cells and also released into the blood from mast cells activated by an injury. Among the heparin molecules is a pentaglycan sequence... 

Due to the lowered level of antithrombin 111 found in some families , or during treatment with heparin, it has been suggested that the intravenous administration of purified anti-thrombin III would be beneficial... 

L. Gugliotta, A. D Angelo, M. Mattioli-Belmonte, et al. Hypercoagulability during 1-asparaginase treatment the effect of anti thrombin III supplementation in vivo. Br. J. Haematol. 74 465 (1990). 

Anti thrombin III, an anticoagulant and antithrombotic agent (50 to 100 lU/min IV), is indicated for prophylaxis and adjunct treatment of thromboembolism associated with hereditary antithrombin III deficiency (see also Tables 17 and 18). 

A series of experiments were made with heparin-PVA bead columns. Thrombin (100 U crude bovine, II50 U purified human or 100 U purified bovine) was loaded onto the columns and the excess removed with PBS. Antithrombin III ( 2000-2500 U purified human) was then loaded, and the columns eluted with PBS, (w/v) bovine serum albumin, or plasma fractions. In some cases heat defibrinated or arvinized plasma was used instead of purified human antithrombin III as a crude source of antithrombin III, as well as a displacing agent. The inactivation of thrombin by anti thrombin III was shown in a separate experiment using the chromogenic substrate assay. 

Analysis of the radioactivity displaced by arvinized plasma indicated the presence of thrombin-anti thrombin III complex (- 70%) and what was presumed to be thrombin-a-2-macroglobulin complex (" 30%). The bound thrombin is thought to react first with antithrombin III to produce a bound inactivated thrombin-anti thrombin III complex, which is dislodged from heparin by a yet unknown plasma component(s), decomplexed by an unknown mechanism to react with a-2-macroglobulin. This mechanism is illustrated in Figure 1. After displacement, the increase in I-antithrombin III which had lost its affinity for heparin-Sepharose was attributed to the production of a post complex antithrombin III on decomplexation of the inactive complex. This modified antithrombin III has been described by Lam et al. ( ), Fish et al. (25) and Marciniak (26). Neither free I-thrombin nor I-antithrombin III were detected in the displaced eluent. 

Although displacement of I-thrombin was limited, some heparin sites were regenerated. Further experiments, not yet published (27)> have demonstrated that the immobilized heparin appeared to retain its ability to accelerate the inactivation of thrombin at least over 10 cycles of exposure to thrombin and anti thrombin III. This suggested that the regenerated heparin sites can retain their catalytic... 

The covalent modification of human a-thrombin with pyridoxal 5 -phosphate involves phosphopyridoxylation at two sites.Attachment of the cofactor to the first site was accompanied by a loss in the fibrinogen clotting activity of the thrombin, whereas attachment of the cofactor to the second site resulted in a decrease in the sensitivity to heparin in the anti-(thrombin III)-thrombin reaction. 

Anti-(soluble egg antigen)antibody Anti-tetanolysin antibody Anti-thrombin III antibody 4-Azobenzeneaisonate ... 

We now report results concerning the interactions between these two proteins and two of these polystyrene derivatives sulfonated polystyrene (PSSO3) and sulfonate-glutamic acid sulfonamide polystyrene (PSSO2GIU). The adsorption of each protein was studied either in purified system or in plasma and the binding constants were determined. Moreover, the catalysed formation of thrombin-anti-thrombin III complex was examined and compared with the antithrombic activity measured through thrombin clotting times of plasma preincubated with these polymers. 

Heparin fractions of different molecular weights and anticoagulant activities have been prepared by affinity chromatography on immobilized protamine. A heparin fraction (mol. wt. 7.9 x lO ) exhibited binding to anti-thrombin III with a stoicheiometry of 2 1, whereas two higher molecular weight fractions exhibited 1 1 binding. Similar results have been reported for two fractions from porcine... 

Human anti-thrombin III contains residues of D-galactose, D-mannose, 2-acetamido-2-deoxy-D-glucose, and neuraminic acid. The neuraminic acid residues do not appear to be required for the inhibitory action of this glycoprotein on thrombin, and therefore their presence in the glycoprotein may delay the clearance of the glycoprotein from the bloodstream, as has been shown for ceruloplasmin and ai-acid glycoprotein. 

Heparin continues to be used as a ligand and a wide variety of proteins may be purified on affinity columns prepared from this material. Perhaps its most important use is in the purification of proteins involved in, or associated with, the complex process of blood-clotting e.g. anti-thrombin III, platelet factor 4, factor XI and factor XII ). Other proteins purified using heparin include an oestradiol receptor, arginine-rich high density lipoprotein as well as a number... 

In studies on human anti-thrombin, jS-D-glucosylceramide was found to complex firmly with anti-thrombin III. It was suggested that the antithrombin might act as a carrier for glycolipids. 

Figure 14.3. ES mass spectra of anti-thrombin III acquired under denaturing conditions of progressively increasing severity in moving from top to bottom In addition to the native conformation, at least three distinct non-native states are revealed. Peaks labeled with circles indicate impurities. (Reprinted from Ref. 63 with permission.)...

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