STRUCTURAL STUDIES ON SERINE PROTEASES

University Of Chicago, Chicago IL

Investigators

Linked publications, trials & patents

Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. This proposal focuses on biologically relevant serine proteases in complex with inhibitors, substrates and effectors for which we currently lack structural information. The focus is mainly on thrombin and activated protein C, two key proteases involved in the progression (thrombin) and inhibition (activated protein C) of blood coagulation. We plan to address a number of unsolved issues that will benefit tremendously from the availability of crystal structures. In the case of thrombin, we plan to crystallize the enzyme in complex with thrombomodulin and protein C to establish the molecular basis of its anticoagulant activity. We plan to crystallize the thrombin mutant W215A/E217A, that is entering Phase I clinical trials, in complex with the platelet receptor GPIb to identify the mode of binding underscoring this newly discovered interaction. We plan to crystallize prothrombin in order to obtain information on the architecture of the zymogen form of thrombin. We also plan to crystallize meizothrombin, the most relevant intermediate along the prothrombin pathway, in complex with GPIb and fragments of the protease activated receptors PAR1, PAR3 and PAR4, thrombomodulin and protein C. In the case of activated protein C, we plan to crystallize the enzyme in the E*, E and E:Na+ forms and in complex with fragments of PAR1 and coagulation factor Va. Structures of these proteins and their complexes will produce major advances in our understanding of the moelcular basis of thrombin procoagulant, prothrombotic and anticoagulant activities in the blood, and the way the enzyme is activated from prothrombin via meizothrombin. The structure of W215A/E217A bound to GPIb will reveal the molecular basis of the remarkable antithrombotic effect of this clinically relevant mutant. Structures of activated protein C will advance our understanding of the function of this enzyme in the control of thrombosis and inflammation.