Heparin has been used clinically for over forty years for the treatment of deep vein thrombosis, pulmonary embolism, and other thrombotic disorders in hospitalized patients. However, while the clinical effectiveness of heparin is well established, a complete description of how heparin works in vivo remains to be determined. The long term objective of the research described in this application is to identify effects of heparin on the process of coagulation which are involved in the anticoagulant and antithrombotic mechanisms of action of heparin and to quantitatively relate these effects to the physical-chemical (structural) properties of heparin to ultimately provide a rational basis for heparin therapy and for the preparation of safer synthetic heparin-like drugs. At the present time, it is clear that heparin greatly accelerates the rates at which most of the serine proteases formed during the process of coagulation are inhibited by antithrombin III. Relatively little quantitative data is available, however, on how protein-protein and protein-phosphoslipid interactions, which occur during coagulation, affect the heparin-enhanced antithrombin III/protease reactions.
The specific aim of the research described in this application is to rigorously study the effects of heparin and heparin-antithrombin III on the rate of factor X activation by factor IXa in the presence and absence of cofactors (calcium, phospholipid, factor VIII), to determine (1) how protein-protein and protein-phospholipid interactions in this model system affect heparin-antithrombin III/protease reactions and (2) how these interactions are affected by heparin alone. Antithrombin III reaction rates with factors IXa and Xa will be measured in the presence and absence of cofactors and heparin to determine the kinetic mechanism of action of heparin in these reactions and to determine how various combinations of cofactors affect the reaction rates. The rates of factor X activation by factor IXa in the presence of various combinations of cofactors and heparin will be measured both in the absence and presence of antithrombin III.
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