Selective Loss of Fibrinogen Clotting in a Loop-less Thrombin

Dang, Quoc D.; Sabetta, Michela; Di, Enrico

doi:10.1074/jbc.272.32.19649

Cited by 46 publications

(94 citation statements)

References 19 publications

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“…9,10,[38][39][40] Dang et al prepared an ␣-IIa variant lacking the ␥-loop ("loopless") and noted a 240-fold reduction in fibrinogen cleavage compared with ␣-IIa. 41 Based on an analysis of crystal structures, Soslau et al proposed that conversion of ␣-IIa to ␤-IIa and then ␥-IIa is accompanied by a progressive opening of the active site to take on a more trypsin-like conformation. 39 Physiologic roles for ␤-IIa and ␥-IIa have not been established, but several possibilities have been discussed.…”

Section: Discussionmentioning

confidence: 99%

Activation of factor XI by products of prothrombin activation

Matafonov

Sarilla

Sun

et al. 2011

Blood

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The prothrombinase complex converts prothrombin to ␣-thrombin through the intermediate meizothrombin (Mz-IIa). Both ␣-thrombin and Mz-IIa catalyze factor (F) XI activation to FXIa, which sustains ␣-thrombin production through activation of FIX. The interaction with FXI is thought to involve thrombin anion binding exosite (ABE) I. ␣-Thrombin can undergo additional proteolysis to ␤-thrombin and ␥-thrombin, neither of which have an intact ABE I. In a purified protein system, FXI is activated by ␤-thrombin or ␥-thrombin, and by ␣-thrombin in the presence of the ABE I-blocking peptide hirugen, indicating that a fully formed ABE I is not absolutely required for FXI activation. In a FXI-dependent plasma thrombin generation assay, ␤-thrombin, ␥-thrombin, and ␣-thrombins with mutations in ABE I are approximately 2-fold more potent initiators of thrombin generation than ␣-thrombin or Mz-IIa, possibly because fibrinogen, which binds to ABE I, competes poorly with FXI for forms of thrombin lacking ABE I. In addition, FXIa can activate factor FXII, which could contribute to thrombin generation through FXIIa-mediated FXI activation. The data indicate that forms of thrombin other than ␣-thrombin contribute directly to feedback activation of FXI in plasma and suggest that FXIa may provide a link between tissue factorinitiated coagulation and the proteases of the contact system. (Blood. 2011;118(2): 437-445) IntroductionThe trypsin-like protease ␣-thrombin (␣-IIa) is a key contributor to vital host responses to injury, including fibrin formation, 1-3 platelet and endothelial cell activation, 4 and inflammation. 5 During coagulation, prothrombin is converted to ␣-IIa through a series of proteolytic steps catalyzed by factor (F) Xa. 6,7 The process results in formation of a functional active site and expression of 2 anion binding exosites (ABE I and ABE II) that are required for ␣-IIa interactions with many substrates, receptors, and inhibitors (Table 1). [1][2][3]6 In the presence of FVa and phospholipid, FXa initially converts prothrombin to the protease meizothrombin (Mz-IIa; Figure 1A), 7-11 which expresses ABE I. 6 Mz-IIa is rapidly converted to ␣-IIa, which may undergo further proteolysis to form ␤-thrombin (␤-IIa) and ␥-thrombin (␥-IIa) ( Figure 1B), 2 proteases with reduced capacity to catalyze ABE I-dependent reactions. [9][10][11][12] Physiologic roles for ␤-IIa or ␥-IIa have not been established; however, both have been identified in clotting blood. 11 ␣-IIa up-regulates its own generation by activating the cofactors FV and FVIII, [1][2][3]9 and by converting FXI to the protease FXIa. 13,14 FXIa, in turn, sustains ␣-IIa generation by converting FIX to FIXa␤, 15 and possibly by activating FV and FVIII. 16 In the original cascade/ waterfall hypotheses of coagulation, FXI is activated by FXIIa 17,18 ; however, current models deemphasize this reaction based on the observation that FXI deficiency is associated with abnormal bleeding, whereas FXII deficiency is not. 18 FXI activation by ␣-IIa would explain the phenotypic di...

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Section: Discussionmentioning

confidence: 99%

Activation of factor XI by products of prothrombin activation

Matafonov

Sarilla

Sun

et al. 2011

Blood

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“…There are no differences in the residues involving the active site, the thrombin ␤-insertion loop (also called the Trp 60D loop), or the allosteric Na ϩ -binding site. The minor differences that do exist between species are not anticipated to interfere with interactions of the substrate peptides at the thrombin active site surface (32).…”

Section: Methodsmentioning

confidence: 99%

“…FXIII AP-(28 -41), PAR1-(32-45), and PPACK each contain a proline at the P 2 position. X-ray crystal studies of PAR1- (32)(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45) and PPACK reveal that thrombin contains a hydrophobic surface that can accommodate this proline (9,10). Fbg A␣- (7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20), by contrast, contains a Val at the P 2 position instead of the Pro.…”

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confidence: 99%

“…Table I shows sequences for Fibrinogen A␣ (amino acids 7-20) (Fbg A␣- (7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)), Factor XIII activation peptide (aa 28 -41) (FXIII AP-(28 -41)), D-Phe-Pro-Arg-chloromethylketone (PPACK), and thrombin receptor (aa 32-45) (PAR1- (32)(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45)). In each peptide, there is an Arg residue at the P 1 position 2 that binds within the catalytic cleft of thrombin forming a salt bridge with thrombin amino acid 3 Asp 189 .…”

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confidence: 99%

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Examining Thrombin Hydrolysis of the Factor XIII Activation Peptide Segment Leads to a Proposal for Explaining the Cardioprotective Effects Observed with the Factor XIII V34L Mutation

Trumbo

Maurer

2000

Journal of Biological Chemistry

130

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In the blood coagulation cascade, thrombin cleaves fibrinopeptides A and B from fibrinogen revealing sites for fibrin polymerization that lead to insoluble clot formation. Factor XIII stabilizes this clot by catalyzing the formation of intermolecular cross-links in the fibrin network. Thrombin activates the Factor XIII a 2 dimer by cleaving the Factor XIII activation peptide segment at the Arg 37 -Gly 38 peptide bond. Using a high performance liquid chromatography assay, the kinetic constants K m , k cat , and k cat /K m were determined for thrombin hydrolysis of fibrinogen A␣-(7-20), Factor XIII activation peptide-(28 -41), and Factor XIII activation peptide-(28 -41) with a Val 34 to Leu substitution. This Val to Leu mutation has been correlated with protection from myocardial infarction. In the absence of fibrin, the Factor XIII activation peptide-(28 -41) exhibits a 10-fold lower k cat /K m value than fibrinogen A␣-(7-20). With the Factor XIII V34L mutation, decreases in K m and increases in k cat produce a 6-fold increase in k cat /K m relative to the wild-type Factor XIII sequence. A review of the x-ray crystal structures of known substrates and inhibitors of thrombin leads to a hypothesis that the new Leu generates a peptide with more extensive interactions with the surface of thrombin. As a result, the Factor XIII V34L is proposed to be susceptible to wasteful conversion of zymogen to activated enzyme. Premature depletion may provide cardioprotective effects.Fibrinogen is composed of three chains A␣, B␤, and ␥ arranged into the dimer (A␣B␤␥) 2 . In blood coagulation, the serine protease thrombin cleaves the N-terminal portions of the A␣ and B␤ chains. For the A␣ chain, cleavage occurs at the Arg 16 -Gly 17 peptide bond and fibrinopeptide A (FpA) 1 is released; whereas, for the B␤ chain, cleavage occurs at the Arg 14 -Gly 15 peptide bond and fibrinopeptide B (FpB) is released. Removal of the fibrinopeptides leads to exposure of fibrin polymerization sites that react to form an insoluble blood clot (reviewed in Ref. 1).Activated Factor XIII helps stabilize this clot structure by catalyzing the formation of intermolecular ␥-glutamyl-⑀-lysine cross-links in the fibrin network and in fibrin-enzyme complexes. Factor XIII is a member of a family of enzymes known as transglutaminases that have a catalytic triad, similar to cysteine proteases, composed of amino acids Cys 314 , His 373, and Asp 396 . In plasma, Factor XIII is expressed as a zymogen of the form a 2 b 2 . In the presence of thrombin and calcium, the a 2 unit is released and activated. By contrast, platelet Factor XIII is expressed as the zymogen a 2 unit (reviewed in Ref. 2).The Factor XIII a 2 dimer contains in the N-terminal portion of each monomer a sequence known as the activation peptide (3, 4). Each activation peptide segment crosses the dimer interface and extends over the catalytic site of the opposing Factor XIII a subunit. Cleavage of the activation peptide segments by thrombin at the Arg 37 -Gly 38 peptide bond aids in exposure of the Fact...

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“…The 60-insertion (or ␤-insertion) loop functions like a lid regulating entrance into the active site, and the autolysis (or ␥) loop participates in controlling substrate specificity. Removal of the autolysis loop leads to a thrombin mutant with greatly hindered ability to convert fibrinogen to fibrin (4). In addition to the ␤-and ␥-loops, thrombin exosites ABE I and ABE II also play key roles in regulating substrate specificity (Fig.…”

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confidence: 99%

Ligand Binding to Anion-binding Exosites Regulates Conformational Properties of Thrombin

Malovichko

Sabo²,

Maurer

2013

Journal of Biological Chemistry

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Background: Thrombin utilizes active site and anion-binding exosites (ABE) to fulfill diverse roles. Results: ABE I ligands PAR3(44 -56), PAR1(49 -62), and Hirudin(54 -65) exert different effects on thrombin exosite-active site and exosite-exosite interactions. More consequences occur with added PPACK and ABE II ligands. Conclusion: Thrombin employs ligands to transmit unique events across the enzyme. Significance: Ligand binding may be tailored to therapeutically regulate multifunctional thrombin.

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Selective Loss of Fibrinogen Clotting in a Loop-less Thrombin

Cited by 46 publications

References 19 publications

Activation of factor XI by products of prothrombin activation

Activation of factor XI by products of prothrombin activation

Examining Thrombin Hydrolysis of the Factor XIII Activation Peptide Segment Leads to a Proposal for Explaining the Cardioprotective Effects Observed with the Factor XIII V34L Mutation

Ligand Binding to Anion-binding Exosites Regulates Conformational Properties of Thrombin

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