title: Methylcobalamin: coenzyme M methyltransferase isoenzymes MtaA and MtbA from Methanosarcina barkeri. Cloning, sequencing and differential transcription of the encoding genes, and functional overexpression of the mtaA gene in Escherichia coli.
abstract: Methanosarcina barkeri is known to contain two methyltransferase isoenzymes, here designated MtaA and MtbA, which catalyze the formation of methyl-coenzyme M from methylcobalamin and coenzyme M. The genes encoding the two soluble 34-kDa proteins have been cloned and sequenced. mtaA and mtbA wee found to be located in different parts of the genome, each forming a monocystronic transcription unit. Northern blot analysis revealed that mtaA is preferentially transcribed when M. barkeri is grown on methanol and the mtbA gene when the organism is grown on H2/CO2 or trimethylamine. Comparison of the deduced amino acid sequences revealed the sequences of the two isoenzymes to be 37% identical. Both isoenzymes showed sequence similarity to uroporphyrinogen III decarboxylase from Escherichia coli. The mtaA gene was tagged with a sequence encoding six His placed bp before the mtaA start codon, and was functionally overexpressed in E. coli. 25% of the E. coli protein was found to be active methyltransferase which could be purified in two steps to apparent homogeneity with a 70% yield.
authors.affiliation: Max-Planck-Institut für terrestrische Mikrobiologie und Laboratorium für Mikrobiologie des Fachbereichs Biologie der Philipps-Universität, Marburg, Germany.
authors.forename: U
authors.initials: U
authors.lastname: Harms
authors.forename: R K
authors.initials: RK
authors.lastname: Thauer
mesh_terms: D000595:Amino Acid Sequence; D019843:Archaeal Proteins; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D015246:Deoxyribonuclease EcoRI; D015252:Deoxyribonucleases, Type II Site-Specific; D004926:Escherichia coli / Q000235:genetics; D007527:Isoenzymes / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D017021:Methanosarcina barkeri / Q000201:enzymology / Q000235:genetics*; D008780:Methyltransferases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic
chemical_list: D019843:Archaeal Proteins; D004269:DNA, Bacterial; D007527:Isoenzymes; D008780:Methyltransferases; C023894:methylcobalamin-coenzyme M methyltransferase; D015246:Deoxyribonuclease EcoRI; D015252:Deoxyribonucleases, Type II Site-Specific; C024405:GATC-specific type II deoxyribonucleases