ABSTRACT
The use of a polymer electrolyte fuel cell (PEFC) with a Nafion membrane for isotopic separation of deuterium (D) was investigated. Mass analysis at the cathode side indicated that D diffused through the membrane and participated in an isotope exchange reaction. The exchange of D with protium (H) in H2O was facilitated by a Pt catalyst. The anodic data showed that the separation efficiency was dependent on the D concentration in the source gas, whereby the water produced during the operation of the PEFC was more enriched in D as the D concentration of the source gas was increased.
ABSTRACT
In the final process of blood coagulation, fibrin molecules are stabilized via a catalytic reaction by Factor XIIIA (FXIIIA), a member of the transglutaminase (TGase) 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 TGase 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.
Subject(s)
Factor XIII/chemistry , Fish Proteins/chemistry , Amino Acid Sequence , Animals , CRISPR-Cas Systems , Conserved Sequence , Factor XIII/genetics , Factor XIII/metabolism , Fish Proteins/genetics , Fish Proteins/metabolism , Gene Expression , Genetic Association Studies , Humans , Mutation, Missense , Organ Specificity , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Sequence Deletion , Sf9 Cells , SpodopteraABSTRACT
Calcium-dependent transglutaminases (TGs) are a family of enzymes that catalyze protein cross-linking and/or attachment of primary amines in a variety of organisms. Mammalian TGs are implicated in multiple biological events such as skin formation, blood coagulation, and extracellular matrix stabilization. Medaka (Oryzias latipes) has been used as a model fish to investigate the physiological functions of mammalian proteins. By analysis of the medaka genome, we found seven TGs orthologues, some of which apparently corresponded to the mammalian TG isozymes, TG1, TG2, and Factor XIII. All orthologues had preserved amino acid residues essential for enzymatic activity in their deduced primary structures. In this study, we analyzed biochemical properties of two orthologues (OlTGK1 and OlTGK2) of mammalian epithelium-specific TG (TG1) that are significantly expressed at the transcriptional level. Using purified recombinant proteins for OlTGK1 and OlTGK2, we characterized their catalytic reactions. Furthermore, immunohistochemical analyses of fish sections revealed higher expression in the pancreas (OTGK1), intervertebral disk (OlTGK2) and pharyngeal teeth (OlTGK2) as well as in the skin epidermis.
