Factor XIIIa-derived peptides inhibit transglutaminase activity. Localization of substrate recognition sites

K E Achyuthan, T F Slaughter, M A Santiago, J J Enghild, C S Greenberg

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    Abstract

    Factor XIIIa is a transglutaminase that catalyzes intermolecular gamma-glutamyl-epsilon-lysyl bonds between fibrin and other proteins involved in hemostasis. We synthesized 25 peptides from various regions of factor XIIIa and studied their effects on cross-linking fibrin, N,N'-dimethylcasein, or fibronectin. We found that two peptides, Asn72-Asp97 (peptide-4) and Asp190-Phe230 (peptide-7), inhibited factor XIIIa cross-linking of these substrates. The other peptides did not inhibit factor XIIIa activity. The inhibition of cross-linking was reversed by excess substrate, indicating that the peptides were interacting with fibrin and not factor XIIIa. The peptides were not pseudosubstrates since they were not cross-linked to fibrin. The peptides did not modify the primary amine binding site as increasing the primary amine concentration did not reverse inhibition. Peptides-4 and -7 also had no effect on exposure of the active site of factor XIIIa and no synergistic inhibitory effects were detected. Peptides-4 and -7 had no effect on factor XIIIa binding to fibrin suggesting that the binding sites and the substrate recognition sites were distinct. Synthetic peptides containing shorter amino acid sequences of peptide-4 were inactive. In contrast, the amino-terminal (Asp190-Lys199, Tyr194-Tyr204) and the carboxyl-terminal (Lys221-Phe230) portions of peptide-7 were 20-60-fold less inhibitory compared to intact peptide-7. Peptides-4 and -7 also inhibited guinea pig liver tissue transglutaminase from cross-linking fibrinogen, N,N'-dimethylcasein, and fibronectin. In conclusion, we have identified two regions outside the active site pocket which are important for substrate recognition in factor XIIIa and tissue transglutaminase.
    Original languageEnglish
    JournalJournal of Biological Chemistry
    Volume268
    Issue28
    Pages (from-to)21284-92
    Number of pages8
    ISSN0021-9258
    Publication statusPublished - 1993

    Keywords

    • Amines
    • Amino Acid Sequence
    • Animals
    • Binding Sites
    • Biotin
    • Caseins
    • Cross-Linking Reagents
    • Fibrin
    • Guinea Pigs
    • Humans
    • Liver
    • Molecular Probes
    • Molecular Sequence Data
    • Peptide Fragments
    • Substrate Specificity
    • Transglutaminases

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