Insights into the molecular inactivation mechanism of human activated thrombin-activatable fibrinolysis inhibitor.
SUMMARY BACKGROUND: Thrombin-activatable fibrinolysis inhibitor (TAFI) is a validated target for thrombotic diseases. TAFI is converted in vivo to activated TAFI (TAFIa) by removal of its pro-domain. Whereas TAFI is stable and persists in the circulation, possibly in complex with plasminogen, TAFIa is unstable and poorly soluble, with a half-life of minutes. OBJECTIVES: In order to study the molecular determinants of this instability, we studied the influence of protein inhibitors on human TAFIa. RESULTS: We found that protein inhibitors significantly reduced the instability and insolubility of TAFIa. In addition, we solved the 2.5-A resolution crystal structure of human TAFIa in complex with a potent protein inhibitor, tick-derived carboxypeptidase inhibitor, which gives rise to a stable and soluble TAFIa species. The structure revealed a significant reduction in the flexibility of dynamic segments when compared with the structures of bovine and human TAFI. We also identified two latent hotspots, loop Lbeta2beta3 and segment alpha5-Lalpha5beta7-beta7, where conformational destabilization may begin. These hotspots are also present in TAFI, but the pro-domain may provide sufficient stabilization and solubility to guarantee protein persistence in vivo. When the pro-domain is removed, the free TAFIa moiety becomes unstable, its activity is suppressed, and the molecule becomes insoluble. CONCLUSIONS: The present study corroborates the function of protein inhibitors in stabilizing human TAFIa and it provides a rigid and high-resolution mold for the design of small molecule inhibitors of this enzyme, thus paving the way for novel therapy for thrombotic disorders.
Sanglas, L; Arolas, JL; Valnickova, Z; Aviles, FX; Enghild, JJ; Gomis-Rüth, FX
Volume / Issue
Start / End Page
Pubmed Central ID
Electronic International Standard Serial Number (EISSN)
International Standard Serial Number (ISSN)
Digital Object Identifier (DOI)