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Biochemical characterization of a medaka ( Oryzias latipes ) orthologue for mammalian Factor XIII and establishment of a gene‐edited mutant
Author(s) -
Horimizu Rima,
Ogawa Ryota,
Watanabe Yuko,
Tatsukawa Hideki,
Kinoshita Masato,
Hashimoto Hisashi,
Hitomi Kiyotaka
Publication year - 2017
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.14153
Subject(s) - oryzias , mutant , tissue transglutaminase , biology , thrombin , fibrin , gene , microbiology and biotechnology , proteolysis , recombinant dna , enzyme , biochemistry , platelet , immunology
In the final process of blood coagulation, fibrin molecules are stabilized via a catalytic reaction by Factor XIIIA ( FXIIIA ), a member of the transglutaminase ( TG ase) family that catalyzes protein cross‐linking reactions. In this study, we characterized the orthologue of this enzyme in medaka ( Oryzias latipes ), an established model fish in which a coagulation system is also preserved. The recombinant protein of this orthologue enzyme was produced in baculovirus‐infected insect cells and used for analysis of its biochemical properties including activation by thrombin proteolysis and calcium dependence of the TG ase enzymatic activity. Immunostaining and immunoblotting revealed that medaka FXIIIA is expressed in the kidney, bone, and esophagus in addition to blood cells. Furthermore, a gene‐mutant fish was established using the CRISPR /Cas9 system. The loss of FXIIIA expression was validated in the mutants, and phenotypes, such as absence of fibrin cross‐linking, were investigated in the established mutant fish.