Publications

• Gamma-glutamylmethylamide. A new intermediate in the metabolism of methylamine.

  Kung H.F., Wagner C.

  J Biol Chem 244:4136-4140(1969) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Genetics of the glutamate-mediated methylamine utilization pathway in the facultative methylotrophic beta-proteobacterium Methyloversatilis universalis FAM5.

  Latypova E., Yang S., Wang Y.S., Wang T., Chavkin T.A., Hackett M., Schafer H., Kalyuzhnaya M.G.

  The ability of some microbial species to oxidize monomethylamine via glutamate-mediated pathways was proposed in the 1960s; however, genetic determinants of the pathways have never been described. In the present study we describe a gene cluster essential for operation of the N-methylglutamate path ... >> More

  The ability of some microbial species to oxidize monomethylamine via glutamate-mediated pathways was proposed in the 1960s; however, genetic determinants of the pathways have never been described. In the present study we describe a gene cluster essential for operation of the N-methylglutamate pathway in the methylotrophic beta-proteobacterium Methyloversatilis universalis FAM5. Four major polypeptides from protein fractions displaying high activities of N-methylglutamate synthetase, N-methylglutamate dehydrogenase and gamma-glutamylmethylamide synthetase were selected for mass spectrometry-based identification. The activities of enzymes were associated with the presence of peptides identified as ferredoxin-dependent glutamate synthase (GltB2), large subunit of putative heterotetrameric sarcosine oxidase (SoxA) and glutamine synthetase type III (GSIII) respectively. A gene cluster (8.3 kb) harbouring gltB2, soxA and gsIII-like genes was amplified from M. universalis FAM5, sequenced and assembled. Two partial and six complete open reading frames arranged in the order soxBDAG-gsIII-gltB132 were identified and subjected to mutational analysis, functional and metabolic profiling. We demonstrated that gltB-like and sox-like genes play a key role in methylamine utilization and encode N-methylglutamate synthetase and N-methylglutamate dehydrogenase respectively. Metabolic, enzymatic and mutational analyses showed that the gsIII-like gene encodes gamma-glutamylmethylamide synthetase; however, this enzyme is not essential for oxidation of methylamine. << Less

  Mol. Microbiol. 75:426-439(2010) [PubMed] [EuropePMC]

• Cloning and expression of Methylovorus mays No. 9 gene encoding gamma-glutamylmethylamide synthetase: an enzyme usable in theanine formation by coupling with the alcoholic fermentation system of baker's yeast.

  Yamamoto S., Wakayama M., Tachiki T.

  Gamma-glutamylmethylamide synthetase (GMAS), found in an obligate methylotroph, Methylovorus mays No. 9, can form theanine from glutamic acid and ethylamine in a mixture in which yeast sugar fermentation is coupled for ATP regeneration. The internal and N-terminal amino acid sequences of GMAS had ... >> More

  Gamma-glutamylmethylamide synthetase (GMAS), found in an obligate methylotroph, Methylovorus mays No. 9, can form theanine from glutamic acid and ethylamine in a mixture in which yeast sugar fermentation is coupled for ATP regeneration. The internal and N-terminal amino acid sequences of GMAS had certain similarities to putative glutamine synthetase type III (GS III) of Methylobacillus flagellatus KT. From the M. mays No. 9 genomic DNA library, a clone containing a 6.5-kbp insertional DNA fragment was selected by the PCR screening technique with oligonucleotide primers specific for the GMAS gene. The fragment had an open reading frame of the GMAS gene encoding a protein of 444 amino acids (molecular mass, 49 kDa). The deduced amino acid sequence showed significant identity with that of Met. flagellatus KT GS III (78%). The isolated gene was ligated into an expression vector (pET21a) and expressed in Escherichia coli AD494 (DE3). Enzyme productivity in the expression system was about 23-fold higher than that in M. mays No. 9. Recombinant GMAS had the same properties as intrinsic GMAS, and it formed theanine by coupling the reaction with the ATP-regeneration system of yeast sugar fermentation. << Less

  Biosci. Biotechnol. Biochem. 72:101-109(2008) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Characterization of theanine-forming enzyme from Methylovorus mays no. 9 in respect to utilization of theanine production.

  Yamamoto S., Wakayama M., Tachiki T.

  For development of theanine production from glutamic acid and ethylamine by coupling yeast sugar fermentation as an ATP-regenerating system, several strains were selected from among about 200 methylamine-and/or methanol-assimilating bacteria depending on the theanine-forming activity of their perm ... >> More

  For development of theanine production from glutamic acid and ethylamine by coupling yeast sugar fermentation as an ATP-regenerating system, several strains were selected from among about 200 methylamine-and/or methanol-assimilating bacteria depending on the theanine-forming activity of their permeated cells. The amount of theanine formed by the cells of the selected strains was much larger than that by the cells of Escherichia coli AD494 (DE3) expressing Pseudomonas taetrolens Y-30 glutamine synthetase (GS), which has been found to be a usable enzyme for theanine production. A GS-like enzyme responsible for the theanine-forming reaction was obtained from an obligate methylotroph isolate, Methylovorus mays No. 9. The enzyme was induced by methylamine in the culture medium. A molecular mass of 410-470 kDa was obtained by gel filtration of the enzyme, and 51 kDa by SDS-PAGE analysis. The enzyme showed high activity toward methylamine rather than ammonia, which indicates that it is similar to known gamma-glutamylmethylamide synthetase. The isolated enzyme also had high reactivity to ethylamine in a neutral pH range, and formed theanine from glutamic acid and ethylamine in a reaction mixture containing a yeast sugar fermentation system for ATP-regeneration. << Less

  Biosci. Biotechnol. Biochem. 71:545-552(2007) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.