title: In vitro modification of betaine-homocysteine S-methyltransferase by tissue-type transglutaminase.
abstract: Transglutaminases catalyze the cross-linking and amine incorporation of proteins, and are implicated in various biological phenomena. To elucidate the physiological roles of transglutaminase at the molecular level, we need to identify its physiological protein substrates and clarify the relationship between transglutaminase modification of protein substrates and biological responses. Here we examined whether betaine-homocysteine S-methyltransferase (BHMT: EC 2.1.1.5) can be a substrate of tissue-type transglutaminase by in vitro experiments using porcine liver BHMT and guinea pig liver transglutarninase. Guinea pig liver transglutaminase incorporated 5-(biotinamido) pentylamine and [3H] histamine into BHMT in a time-dependent manner. Putrescine and spermidine also seemed to be incorporated into BHMT by transglutaminase. In the absence of the primary amines, BHMT subunits were cross-linked intra- and intermolecularly. BHMT activity was decreased significantly through the cross-linking by transglutaminase. Histamine incorporation slightly reduced the BHMT activity. Peptide fragments of BHMT containing the glutamine residues reactive for transglutaminase reaction were isolated through biotin labelling, proteinase digestion, biotin-avidin a affinity separation, and reverse phase HPLC. The results of amino acid sequence analyses of these peptides and sequence homology alignment with other mammalian liver BHMT subunits showed that these reactive glutamine residues were located in the region near the carboxyl terminal of porcine BHMT subunit. These results suggested that the liver BHMT can be modified by tissue-type transglutaminase and its activity is regulated repressively by the modification, especially by the cross-linking. This regulatory reaction might be involved in the regulation of homocysteine metabolism in the liver.
authors.affiliation: Department of Applied Biology, Kyoto Institute of Technology, Matsugasaki, Sakyo-ku, Kyoto 606-8585, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Ichikawa
authors.forename: Yuji
authors.initials: Y
authors.lastname: Ohashi
authors.forename: Shinpei
authors.initials: S
authors.lastname: Terada
authors.forename: Shunji
authors.initials: S
authors.lastname: Natsuka
authors.forename: Koji
authors.initials: K
authors.lastname: Ikura
mesh_terms: D000595:Amino Acid Sequence; D000641:Ammonia / Q000378:metabolism; D000818:Animals; D050940:Betaine-Homocysteine S-Methyltransferase; D002850:Chromatography, Gel; D003432:Cross-Linking Reagents; D006168:Guinea Pigs; D008099:Liver / Q000201:enzymology; D008780:Methyltransferases / Q000378:metabolism*; D008969:Molecular Sequence Data; D011503:Transglutaminases / Q000378:metabolism*
chemical_list: D003432:Cross-Linking Reagents; D000641:Ammonia; D008780:Methyltransferases; D050940:Betaine-Homocysteine S-Methyltransferase; D011503:Transglutaminases