UC San Diego

The serine protease thrombin plays an essential regulatory role in blood clotting. Prothrombin is converted to the active α-thrombin in response to tissue damage, allowing the enzyme to cleave procoagulative substrates such as fibrinogen and PAR, enabling the formation of the clot. On the other hand, when thrombin binds its protein cofactor thrombomodulin (TM), the enzyme loses substrate specificity for procoagulative substrates, and engages the anticoagulative pathway by selectively targeting the substrate protein C for proteolytic activation. Despite decades of study, the mechanism by which TM enables thrombin to cleave protein C has yet to be fully characterized, indicating a need for further investigation. Crystallographic evidence shows no notable difference between the thrombin and thrombin-TM structures, suggesting the influence of TM is dynamic in nature. The work presented in this thesis uses two biophysical techniques, hydrogen-deuterium exchange mass spectrometry (HDXMS) and nuclear magnetic resonance spectrometry (NMR) to report the effects of TM on thrombin dynamics.