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Chloromethane-induced genes define a third C1 utilization pathway in Methylobacterium chloromethanicum CM4.

ABSTRACT: Methylobacterium chloromethanicum CM4 is an aerobic alpha-proteobacterium capable of growth with chloromethane as the sole carbon and energy source. Two proteins, CmuA and CmuB, were previously purified and shown to catalyze the dehalogenation of chloromethane and the vitamin B12-mediated transfer of the methyl group of chloromethane to tetrahydrofolate. Three genes located near cmuA and cmuB, designated metF, folD and purU and encoding homologs of methylene tetrahydrofolate (methylene-H4folate) reductase, methylene-H4folate dehydrogenase-methenyl-H4folate cyclohydrolase and formyl-H4folate hydrolase, respectively, suggested the existence of a chloromethane-specific oxidation pathway from methyl-tetrahydrofolate to formate in strain CM4. Hybridization and PCR analysis indicated that these genes were absent in Methylobacterium extorquens AM1, which is unable to grow with chloromethane. Studies with transcriptional xylE fusions demonstrated the chloromethane-dependent expression of these genes. Transcriptional start sites were mapped by primer extension and allowed to define three transcriptional units, each likely comprising several genes, that were specifically expressed during growth of strain CM4 with chloromethane. The DNA sequences of the deduced promoters display a high degree of sequence conservation but differ from the Methylobacterium promoters described thus far. As shown previously for purU, inactivation of the metF gene resulted in a CM4 mutant unable to grow with chloromethane. Methylene-H4folate reductase activity was detected in a cell extract of strain CM4 only in the presence of chloromethane but not in the metF mutant. Taken together, these data provide evidence that M. chloromethanicum CM4 requires a specific set of tetrahydrofolate-dependent enzymes for growth with chloromethane.

SUBMITTER: Studer A

PROVIDER: S-EPMC135114 | biostudies-literature | 2002 Jul

REPOSITORIES: biostudies-literature

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Chloromethane-induced genes define a third C1 utilization pathway in Methylobacterium chloromethanicum CM4.

Studer Alex A McAnulla Craig C Büchele Rainer R Leisinger Thomas T Vuilleumier Stéphane S

Journal of bacteriology 20020701 13

Methylobacterium chloromethanicum CM4 is an aerobic alpha-proteobacterium capable of growth with chloromethane as the sole carbon and energy source. Two proteins, CmuA and CmuB, were previously purified and shown to catalyze the dehalogenation of chloromethane and the vitamin B12-mediated transfer of the methyl group of chloromethane to tetrahydrofolate. Three genes located near cmuA and cmuB, designated metF, folD and purU and encoding homologs of methylene tetrahydrofolate (methylene-H4folate) ...[more]

PMID: 12057941

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Project description:Chloromethane (CH3Cl) is the most abundant volatile halocarbon in the atmosphere and contributes to the destruction of stratospheric ozone. The only known pathway for bacterial chloromethane utilization (cmu) was characterized in Methylobacterium extorquens CM4, a methylotrophic bacterium able to utilize compounds without carbon-carbon bonds such as methanol and chloromethane as the sole carbon source for growth. Previous work demonstrated that tetrahydrofolate and vitamin B12 are essential cofactors of cmuA- and cmuB-encoded methyltransferases of chloromethane dehalogenase, and that the pathway for chloromethane utilization is distinct from that for methanol. This work reports genomic and proteomic data demonstrating that cognate cmu genes are located on the 380 kb pCMU01 plasmid, which drives the previously defined pathway for tetrahydrofolate-mediated chloromethane dehalogenation. Comparison of complete genome sequences of strain CM4 and that of four other M. extorquens strains unable to grow with chloromethane showed that plasmid pCMU01 harbors unique genes without homologs in the compared genomes (bluB2, btuB, cobA, cbiD), as well as 13 duplicated genes with homologs of chromosome-borne genes involved in vitamin B12-associated biosynthesis and transport, or in tetrahydrofolate-dependent metabolism (folC2). In addition, the presence of both chromosomal and plasmid-borne genes for corrinoid salvaging pathways may ensure corrinoid coenzyme supply in challenging environments. Proteomes of M. extorquens CM4 grown with one-carbon substrates chloromethane and methanol were compared. Of the 49 proteins with differential abundance identified, only five (CmuA, CmuB, PurU, CobH2 and a PaaE-like uncharacterized putative oxidoreductase) are encoded by the pCMU01 plasmid. The mainly chromosome-encoded response to chloromethane involves gene clusters associated with oxidative stress, production of reducing equivalents (PntAA, Nuo complex), conversion of tetrahydrofolate-bound one-carbon units, and central metabolism. The mosaic organization of plasmid pCMU01 and the clustering of genes coding for dehalogenase enzymes and for biosynthesis of associated cofactors suggests a history of gene acquisition related to chloromethane utilization.

Dataset Information

Chloromethane-induced genes define a third C1 utilization pathway in Methylobacterium chloromethanicum CM4.

Publications

Chloromethane-induced genes define a third C1 utilization pathway in Methylobacterium chloromethanicum CM4.

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