Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.5 (thrombin)
 33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Coagulation factor IX is a vitamin K-dependent glycoprotein that circulates in blood as a precursor of a serine protease. Incubation of human factor IX with human alpha-thrombin resulted in a time and enzyme concentration-dependent cleavage of factor IX yielding a molecule composed of a heavy chain (mol wt 50,000) and a doublet light chain (mol wt 10,000). The proteolysis of factor IX by thrombin was significantly inhibited by physiological levels of calcium ions. Under nondenaturing conditions, the heavy and light chains of thrombin-cleaved factor IX remained strongly associated, but these chains were readily separated by gel filtration in the presence of denaturants. Amino-terminal sequence analyses of the isolated heavy and light chains of thrombin-cleaved human factor IX indicated that thrombin cleaved peptide bonds at Arg327-Val328 and Arg338-Ser339 in this molecule. Comparable cleavages were observed in bovine factor IX by bovine thrombin and occurred at Arg319-Ser320 and Arg339-Ser340. Essentially, a complete loss of factor IX procoagulant activity was associated with its cleavage by thrombin. Furthermore, thrombin-cleaved factor IX neither developed coagulant activity after treatment with factor XIa nor inhibited the coagulant activity of native factor IX. These data indicate that thrombin cleaves factor IX near its active site serine residue, rendering it incapable of activating factor X. Whether or not this reaction occurs in vivo is unknown.
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PMID:Proteolytic inactivation of blood coagulation factor IX by thrombin. 406 23

Coagulation factor IX has the modular composition Gla-(EGF)2-SP, where Gla, EGF, and SP represent the gamma-carboxy-Glu-rich, epidermal growth factor-like, and serine protease modules, respectively. The protein melts in two distinct temperature regions. The SP module melts at lower temperature between 42 and 55 degrees C, depending on the pH, with irreversible loss of esterolytic activity. The endotherm corresponding to this transition is readily described by a two-state transition indicating the melting of a single cooperative unit. A thrombin-generated 12-kDa fragment representing the COOH-terminal half of the SP module and a 45-kDa fragment containing the NH2-terminal half of the SP module and the rest of the molecule can be separated by exclusion chromatography in 3 M urea and recombined in its absence. Both fragments retain a compact structure as evidenced by melting transitions near 60 degrees C at neutral pH. This indicates that the SP module contains two independently folded domains that strongly interact with each other and seem to merge into one cooperative unit in the intact protein. At low pH at high temperature a second peak appears which is also observed in a 19-kDa fragment containing the EGF modules. Thermodynamic analysis of this second peak showed that the two EGF modules are independently folded and provided evidence for a weak interaction between them. Fluorescence and CD measurements indicated that the secondary structure of the isolated 6-kDa Gla fragment is substantially increased in the presence of Ca2+. The Ca2+-loaded Gla fragment undergoes a sigmoidal unfolding transition as revealed by fluorescence and CD measurements. This same transition in a 25-kDa Gla-(EGF)2 fragment was stabilized by more than 10 degrees C, indicating a strong interaction between the Ca(2+)-loaded Gla and EGF domains. Thus, factor IX consists of five independently folded interacting domains.
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PMID:Domain structure and domain-domain interactions in human coagulation factor IX. 847 87