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Thromb Haemost 2020; 120(03): 400-411
DOI: 10.1055/s-0039-3402760
Coagulation and Fibrinolysis
Georg Thieme Verlag KG Stuttgart · New York

The Fibronectin Type II Domain of Factor XII Ensures Zymogen Quiescence

Chantal C. Clark‡   *
1   Department of Clinical Chemistry and Haematology, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands
,
Zonne L. M. Hofman‡   *
1   Department of Clinical Chemistry and Haematology, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands
2   The Laboratory for Translational Immunology, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands
,
Wariya Sanrattana
1   Department of Clinical Chemistry and Haematology, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands
,
Lyanne den Braven
1   Department of Clinical Chemistry and Haematology, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands
,
Steven de Maat
1   Department of Clinical Chemistry and Haematology, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands
,
Coen Maas
1   Department of Clinical Chemistry and Haematology, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands
› Institutsangaben
Funding C.M. gratefully acknowledges financial support from the Netherlands Thrombosis Foundation and Landsteiner Foundation for Blood Transfusion Research. Z.I.M.H. is supported by an Alexandre Suermann Fellowship from the Utrecht University Medical Center. W.S. gratefully acknowledges financial support from the Royal Thai Government.
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Publikationsverlauf

26. Juni 2019

27. November 2019

Publikationsdatum:
15. Januar 2020 (online)

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Abstract

Factor XII (FXII) zymogen activation requires cleavage after arginine 353 located in the activation loop. This cleavage can be executed by activated FXII (autoactivation), plasma kallikrein (PKa), or plasmin. Previous studies proposed that the activation loop of FXII is shielded to regulate FXII activation and subsequent contact activation. In this study, we aimed to elucidate this mechanism by expressing and characterizing seven consecutive N-terminally truncated FXII variants as well as full-length wild-type (WT) FXII. As soon as the fibronectin type II domain is lacking (FXII Δ1–71), FXII cleavage products appear on Western blot. These fragments display spontaneous amidolytic activity, indicating that FXII without the fibronectin type II domain is susceptible to autoactivation. Additionally, truncated FXII Δ1–71 is more easily activated by PKa or plasmin than full-length WT FXII. To exclude a contribution of autoactivation, we expressed active-site incapacitated FXII truncation variants (S544A). FXII S544A Δ1–71 is highly susceptible to cleavage by PKa, indicating exposure of the activation loop. In surface binding experiments, we found that the fibronectin type II domain is non-essential for binding to kaolin or polyphosphate, whereas the following epidermal growth factor-like domain is indispensable. Binding of full-length FXII S544A to kaolin or polyphosphate increases its susceptibility to cleavage by PKa. Moreover, the activation of full-length WT FXII by PKa increases approximately threefold in the presence of kaolin. Deletion of the fibronectin type II domain eliminates this effect. Combined, these findings suggest that the fibronectin type II domain shields the activation loop of FXII, ensuring zymogen quiescence.

Keywords

factor XII - plasma kallikrein - bradykinin - contact system - fibronectin type II domain - epidermal growth factor-like domain - polyphosphate - plasmin

Authors' Contributions

C.C.C., Z.L.M.H., W.S., S.d.M., and C.M. performed experiments, were involved in the development of the concept, design and interpretation of data, and wrote the manuscript.


* Authors contributed equally.


Supplementary Material