A model for the unique role of factor Va A2 domain extension in the human ternary thrombin-generating complex

Shim, Joong‐Youn; Lee, Chang Jun; Wu, Sangwook; Pedersen, Lee G.

doi:10.1016/j.bpc.2015.02.003

Cited by 18 publications

(45 citation statements)

References 21 publications

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“…Consequently, the latter was generated by Huntington and coworkers . More recently, a full human ternary complex was reported, thereby providing important new insights in the structural organization of the prothrombinase complex and prothrombin binding. In the following sections, we will review the molecular details of the FVa‐FXa interactions, guided by the crystal structure of pseutarin C, structural homology models, and biochemical studies.…”

Section: The Factor Va‐factor Xa Interactionmentioning

confidence: 99%

Blood coagulation factor Va's key interactive residues and regions for prothrombinase assembly and prothrombin binding

Schreuder

Reitsma

Bos

2019

Journal of Thrombosis and Haemostasis

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Blood coagulation factor Va serves an indispensable role in hemostasis as cofactor for the serine protease factor Xa. In the presence of an anionic phospholipid membrane and calcium ions, factors Va and Xa assemble into the prothrombinase complex. Following formation of the ternary complex with the macromolecular zymogen substrate prothrombin, the latter is rapidly converted into thrombin, the key regulatory enzyme of coagulation. Over the years, multiple binding sites have been identified in factor Va that play a role in the interaction of the cofactor with factor Xa, prothrombin, or the anionic phospholipid membrane surface. In this review, an overview of the currently available information on these interactive sites in factor Va is provided, and data from biochemical approaches and 3D structural protein complex models are discussed. The structural models have been generated in recent years and provide novel insights into the molecular requirements for assembly of both the prothrombinase and the ternary prothrombinase‐prothrombin complexes. Integrated knowledge of functionally important regions in factor Va will allow for a better understanding of factor Va cofactor activity.

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Section: The Factor Va‐factor Xa Interactionmentioning

confidence: 99%

Blood coagulation factor Va's key interactive residues and regions for prothrombinase assembly and prothrombin binding

Schreuder

Reitsma

Bos

2019

Journal of Thrombosis and Haemostasis

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“…The significant increase in affinity of prothrombinase for its substrate is attributed to tighter binding of the enzymatic complex to FII because of its localization on the membrane surface by fVa. The longstanding hypothesis that fVa "localizes and positions" FII in an optimum position for efficient catalysis by fXa consistent with the classical role of a cofactor for catalysis was recently confirmed by computational studies with prothrombinase by Shim et al (27). These studies demonstrated that the acidic COOH-terminal portion of the FIGURE 9.…”

Section: Discussionmentioning

confidence: 72%

“…This interaction was further discounted without providing any solid evidence (53) despite initial findings by Guinto and Esmon (54) and more recent original findings from our laboratory (47)(48)(49). A very recent model of prothrombinase using as a template the crystal structure of prothrombinase from the snake venom of Pseudonaja textilis verified and established the critical role of the acidic COOHterminal region of fVa heavy chain for optimal rates of FII cleavage at two spatially distinct sites by prothrombinase resulting in timely IIa formation at the place of vascular injury (27,55).…”

mentioning

confidence: 71%

“…This increase is mainly associated with a large (3,000-fold) increase in the k cat value of fXa within prothrombinase with a 100-fold decrease in the K m value of the enzyme (16). This substantial increase in enzymatic activity resulting in rapid and physiologically relevant IIa generation at the place of vascular injury is credited through precise and unique interactions of the cofactor with specific amino acids affiliated with both membrane-bound fXa and membrane-bound FII as recently demonstrated (27). Accordingly, the introduction of the nonenzymatic cofactor into prothrombinase equips the organism's coagulation artillery necessary for the explosive arrest of vasculature bleeding.…”

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confidence: 99%

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The Dual Regulatory Role of Amino Acids Leu480 and Gln481 of Prothrombin

Wiencek

Hirbawi

Yee

et al. 2016

Journal of Biological Chemistry

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Prothrombin (FII) is activated to ␣-thrombin (IIa) by prothrombinase. Prothrombinase is composed of a catalytic subunit, factor Xa (fXa), and a regulatory subunit, factor Va (fVa), assembled on a membrane surface in the presence of divalent metal ions. We constructed, expressed, and purified several mutated recombinant FII (rFII) molecules within the previously determined fVa-dependent binding site for fXa ( In the presence of a procoagulant membrane surface and divalent metal ions, factor Va (fVa) 3 binds factor Xa (fXa) to form prothrombinase. Prothrombinase is the two-subunit enzymatic complex where the non-enzymatic regulatory subunit (fVa) controls the rate and directs cleavage of prothrombin (FII) by the catalytic subunit (fXa) at two spatially distinct sites resulting in timely ␣-thrombin (IIa) formation at the place of vascular injury (1-3). Cleavage at Arg 271 and Arg 320 of FII is required to form the active serine protease IIa. The essential IIa molecule bears strong homology with other serine protease enzymes, such as activated protein C (APC), chymotrypsin, and fXa. Several different numberings of IIa residues appear in the literature based on either the chymotrypsin numbering (4) or IIa numbering (5, 6) or the entire FII sequence (7). The latter nomenclature is used herein with the appropriate chymotrypsin numbering in parentheses when required for comparison with the existing data in the literature.Historically, it has been shown that in the absence of fVa, initial cleavage at Arg 271 of FII results in the generation of the inactive intermediate prethrombin-2 and fragment 1⅐2. Further cleavage of prethrombin-2 at Arg 320 generates IIa (prethrombin-2 pathway) (8 -21). Concurrent with the appearance of excess fVa during clotting and in the presence of a procoagulant surface, the order of cleavages is reversed, and initial cleavage at Arg 320 generates a transient enzymatically active intermediate, meizothrombin, that has much higher catalytic efficiency than IIa toward chromogenic substrates usually employed to assess IIa activity (18,(22)(23)(24). Meizothrombin is next cleaved at Arg 271 resulting in the generation of IIa and fragment 1⅐2 (meizothrombin pathway). Although efficient cleavage at each site requires the presence of phospholipids, initial cleavage at Arg 320 is entirely fVa-dependent. In the absence of fVa, the two activation cleavage sites are not readily available, and FII is activated at a slow non-physiological rate by membrane-bound fXa alone. Interactions between fXa and FII are known to exist in the presence and absence of fVa; however, the enhanced activity of fXa within prothrombinase toward both activating cleavage sites is controlled solely by the membrane-bound non-enzymatic cofactor (3,16,17,25,26 3 The abbreviations used are: fVa, factor Va; fV, factor V; FII, prothrombin; fXa, factor Xa; IIa, ␣-thrombin; r, recombinant; PC, L-␣-phosphatidylcholine; PCPS, small unilamellar phospholipids vesicles composed of 75% PC and 25% L-␣-phosphatidylserine (w/w); rFII WT , recombinant...

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“…4B) and also as reported in a recent study by Pozzi et al [1], and perhaps play a biological function for prethrombin-2, we examined prethrombin-2 within the context of the processing of prethrombin-2 by prothrombinase. Upon examination for possible contacts of prothrombinase with prothrombin in our recent ternary complex model of prothrombinase-prothrombin [9], we find that R313 and R316 of the FVa A2 domain form tight salt bridges with E186b and D186a, respectively, of the 180s-loop of prothrombin. If we assume that prethrombin-2 and prothrombin are positioned similarly within prothrombinase, as shown in the overlay of prethrombin-2 to prothrombin within prothrombinase of the ternary model ( Supplementary Fig.…”

Section: Ppack-x-ray Alternative-x-ray Collapsed-x-raymentioning

confidence: 98%

Do the crystallographic forms of prethrombin-2 revert to a single form in solution?

Shim

Lee

et al. 2015

Biophysical Chemistry

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A model for the unique role of factor Va A2 domain extension in the human ternary thrombin-generating complex

Cited by 18 publications

References 21 publications

Blood coagulation factor Va's key interactive residues and regions for prothrombinase assembly and prothrombin binding

Blood coagulation factor Va's key interactive residues and regions for prothrombinase assembly and prothrombin binding

The Dual Regulatory Role of Amino Acids Leu480 and Gln481 of Prothrombin

Do the crystallographic forms of prethrombin-2 revert to a single form in solution?

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