Dataset Information

Feedback activation of factor XI by thrombin does not occur in plasma.

ABSTRACT: In this study, we tested the hypothesis that factor XI (FXI) activation occurs in plasma following activation of the extrinsic pathway by thrombin-mediated feedback activation. We used two different assays: (i) a direct measurement of activated FXI by ELISA and (ii) a functional assay that follows the activation of the coagulation cascade in the presence or absence of a FXI inhibiting antibody by monitoring thrombin activity. We failed to detect any FXI activation or functional contribution to the activation of the coagulation cascade in platelet poor or platelet-rich plasma, when activation was initiated by thrombin or tissue factor. Additionally, we found that, in the absence of a contact system inhibitor during blood draw, contact activation of FXI can mistakenly appear as thrombin- or tissue-factor-dependent activation. Thus, activation of FXI by thrombin in solution or on the surface of activated platelets does not appear to play a significant role in a plasma environment. These results call for reevaluation of the physiological role of the contact activation system in blood coagulation.

SUBMITTER: Pedicord DL

PROVIDER: S-EPMC1937556 | biostudies-literature | 2007 Jul

REPOSITORIES: biostudies-literature

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Feedback activation of factor XI by thrombin does not occur in plasma.

Pedicord Donna L DL Seiffert Dietmar D Blat Yuval Y

Proceedings of the National Academy of Sciences of the United States of America 20070725 31

In this study, we tested the hypothesis that factor XI (FXI) activation occurs in plasma following activation of the extrinsic pathway by thrombin-mediated feedback activation. We used two different assays: (i) a direct measurement of activated FXI by ELISA and (ii) a functional assay that follows the activation of the coagulation cascade in the presence or absence of a FXI inhibiting antibody by monitoring thrombin activity. We failed to detect any FXI activation or functional contribution to t ...[more]

PMID: 17652512

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Project description:Essentials Zymogen PK is activated to PKa and cleaves substrates kininogen and FXII contributing to bradykinin generation. Monomeric PKa and dimeric homologue FXI utilize the N-terminal apple domains to recruit substrates. A high-resolution 1.3 Å structure of full-length PKa reveals an active conformation of the protease and apple domains. The PKa protease and four-apple domain disc organization is 180° rotated compared to FXI. SUMMARY: Background Plasma prekallikrein (PK) and factor XI (FXI) are apple domain-containing serine proteases that when activated to PKa and FXIa cleave substrates kininogen, factor XII, and factor IX, respectively, directing plasma coagulation, bradykinin release, inflammation, and thrombosis pathways. Objective To investigate the three-dimensional structure of full-length PKa and perform a comparison with FXI. Methods A series of recombinant full-length PKa and FXI constructs and variants were developed and the crystal structures determined. Results and conclusions A 1.3 Å structure of full-length PKa reveals the protease domain positioned above a disc-shaped assemblage of four apple domains in an active conformation. A comparison with the homologous FXI structure reveals the intramolecular disulfide and structural differences in the apple 4 domain that prevents dimer formation in PK as opposed to FXI. Two latchlike loops (LL1 and LL2) extend from the PKa protease domain to form interactions with the apple 1 and apple 3 domains, respectively. A major unexpected difference in the PKa structure compared to FXI is the 180° disc rotation of the apple domains relative to the protease domain. This results in a switched configuration of the latch loops such that LL2 interacts and buries portions of the apple 3 domain in the FXI zymogen whereas in PKa LL2 interacts with the apple 1 domain. Hydrogen-deuterium exchange mass spectrometry on plasma purified human PK and PKa determined that regions of the apple 3 domain have increased surface exposure in PKa compared to the zymogen PK, suggesting conformational change upon activation.

Dataset Information

Feedback activation of factor XI by thrombin does not occur in plasma.

Publications

Feedback activation of factor XI by thrombin does not occur in plasma.

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