Publications

• Methylcobamide:coenzyme M methyltransferase isozymes from Methanosarcina barkeri: physicochemical characterization, cloning, sequence analysis, and heterologous gene expression.

  Leclerc G.M., Grahame D.A.

  A comparative study was made on the physicochemical characteristics of two isozymes of methylcobamide:-coenzyme M methyltransferase (MT2). Both isozymes catalyzed S-methylation of 2-thioethanesulfonate (coenzyme M) and exhibited similar apparent Km values for coenzyme M of 35 microM (MT2-A) and 20 ... >> More

  A comparative study was made on the physicochemical characteristics of two isozymes of methylcobamide:-coenzyme M methyltransferase (MT2). Both isozymes catalyzed S-methylation of 2-thioethanesulfonate (coenzyme M) and exhibited similar apparent Km values for coenzyme M of 35 microM (MT2-A) and 20 microM (MT2-M). Weak binding to methylcobalamin was indicated by the apparent Km of 14 mM for both isozymes. Cob(I)alamin was established as the major product of the reaction, demonstrating heterolytic cleavage of the methylcobamide carbon-cobalt bond. The isozymes were shown to be zinc-containing metalloproteins. Metal ion chelators strongly inhibited both isozymes. A variety of coenzyme M analogs were tested for activity and/or inhibition. One alternative substrate 3-mercaptopropionate was discovered, with apparent Km 9 mM (MT2-A) and 10 mM (MT2-M). The results suggested an active site geometry in which coenzyme M is bound both by S-coordination to zinc, and electrostatic interaction of the sulfonate with a cationic group on the enzyme. Methanosarcina barkeri genes cmtA and cmtM encoding both isozymes were cloned and sequenced. Both genes encoded proteins with 339 amino acids and predicted molecular masses of 36-37 kDa. Active forms of both isozymes were expressed in Escherichia coli. A conserved segment with the potential for metal binding was found. The possibility of zinc involvement in catalysis of coenzyme M methylation is considered. << Less

  J. Biol. Chem. 271:18725-18731(1996) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• 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.

  Harms U., Thauer R.K.

  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 fo ... >> More

  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. << Less

  Eur. J. Biochem. 235:653-659(1996) [PubMed] [EuropePMC]