Evolutionary adaptation of the γ‐carboxyglutamic acid domain in blood coagulation factor X in the Serpentes suborder

• DOI: 10.1096/fasebj.2021.35.s1.02161
• Publication Date: 2021-06-11T01:17:05Z
• Authors: Shuzhen Cheng; Geraldine Poenou; Ka Cheung; Mark Schreuder; Mettine Bos
• Keywords: Prothrombinase; Factor V

The serine protease factor Xa plays an important role in blood coagulation as it, complex with its cofactor Va, proteolytically activates prothrombin to thrombin, the latter being a key enzyme coagulation. To perform this vital function, undergoes numerous posttranslational modifications including vitamin K-dependent γ-carboxylation of glutamic acid (Glu) residues into so-called Gla residues. A correctly γ-carboxylated GLA domain is essential for interaction negatively charged membrane surface activated platelets or endothelial cells at site injury where clot formation required. Moreover, assembly Va prothrombinase occurs on only. Interestingly, venom some Australian Elapid snakes comprises powerful prothrombin-activating complexes consisting Xa- and Va-like proteins that are specifically expressed gland. We have previously demonstrated capable functioning independently phospholipid membrane. aim uncover putative differential between human snake (Pseudonaja textilis, pt) Xa. do so, we exchanged (hFX) ortholog (ptFX), thereby generating hFX-wt, hFX-ptGLA, ptFX-wt, ptFX-hGLA. FX variants were recombinantly purified employing chromatography techniques. activation by either extrinsic intrinsic tenase was analyzed using peptidyl substrate specific FX. Introduction ptGLA hFX modestly improved catalytic rate reaction, while reaction binding affinity enhanced. Conversely, similar conditions no FXa could be detected following incubation ptFX ptFX-hGLA complex. Assessment FX-specific plasma clotting activity which thrombin-dependent fibrin monitored trigger revealed drastically reduced when hGLA (3% residual activity), unperturbed. Use versus rabbit reagents pathway may explain different observations. exchange resulted overall substantial increase Finally, exchanging did not abolish requirement Va-dependent activation. These data indicate interacts contributes conversion subsequent thrombin formation, point evolutionary adaptation domain.