Purification and properties of formate dehydrogenase from Moraxella sp. strain C-1

Asano, Yasuhisa; Sekigawa, Toshikazu; Inukai, Hisamitsu; Nakazawa, Akiko

doi:10.1128/jb.170.7.3189-3193.1988

Cited by 44 publications

(39 citation statements)

References 28 publications

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“…For each spectrum, 125 scans were collected. The chemical shift for CO 2 is at 125.7 ppm, that for HCO 3 Ϫ is at 161.3 ppm, and that for formate is at 172.1; the signal corresponding to methanol is at 50.1 ppm (not shown).…”

Section: Discussionmentioning

confidence: 99%

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Multiple Formate Dehydrogenase Enzymes in the Facultative Methylotroph Methylobacterium extorquens AM1 Are Dispensable for Growth on Methanol

Chistoserdova¹,

Laukel

Portais

et al. 2004

J Bacteriol

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Formate dehydrogenase has traditionally been assumed to play an essential role in energy generation during growth on C 1 compounds. However, this assumption has not yet been experimentally tested in methylotrophic bacteria. In this study, a whole-genome analysis approach was used to identify three different formate dehydrogenase systems in the facultative methylotroph Methylobacterium extorquens AM1 whose expression is affected by either molybdenum or tungsten. A complete set of single, double, and triple mutants was generated, and their phenotypes were analyzed. The growth phenotypes of the mutants suggest that any one of the three formate dehydrogenases is sufficient to sustain growth of M. extorquens AM1 on formate, while surprisingly, none is required for growth on methanol or methylamine. Further studies of the triple mutant showed that formate was not produced quantitatively and was consumed later in growth. These results demonstrated that all three formate dehydrogenase systems must be inactivated in order to disrupt the formate-oxidizing capacity of the organism but that an alternative formate-consuming capacity exists in the triple mutant.The classic scheme of energy metabolism during methylotrophic growth involves a formate oxidation step except in strains in which all formaldehyde is oxidized in the cyclic ribulose monophosphate cycle (1). Formate dehydrogenase (FDH) activity has been detected in most methylotrophs (3,10,13,15,22,42), and a few FDHs have been purified and analyzed (reviewed in reference 39). In the methylotrophic yeast Candida boidinii, the FDH step was shown not to be essential for methylotrophic growth, but FDH mutants showed reduced growth on methanol (32). However, as the complete C. boidinii genome sequence is not available, the presence of other FDHs is not excluded.Mutant-based analysis of the role of the FDH step in C 1 oxidation has not yet been attempted in methylotrophic bacteria. M. extorquens AM1 offers a convenient model to study this question. It possesses two pathways in which formaldehyde can be oxidized to formate (Fig. 1), one linked to tetrahydromethanopterin (H 4 MPT) and another linked to tetrahydrofolate (H 4 F) (5, 6). The enzymes involved in the two pathways have been studied in detail, and current evidence suggests that the main pathway for oxidizing formaldehyde is the H 4 MPTlinked pathway (reviewed in reference 37). It has been demonstrated recently that this pathway produces formate as an intermediate, a result of a formylmethanofuran transferase/ hydrolase reaction (29), and thus in this pathway one molecule of formate is formed in M. extorquens AM1 per oxidized molecule of a C 1 substrate, such as methanol or methylamine. This formate is subsequently oxidized to CO 2 , presumably by FDH (Fig. 1).An FDH from M. extorquens AM1 has recently been purified and characterized and shown to be a novel, tungsten-containing FDH encoded by two genes, fdh1AB (20). In this study we identified two new regions in the M. extorquens AM1 chromosome coding for two addition...

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Section: Discussionmentioning

confidence: 99%

“…Formate dehydrogenase (FDH) activity has been detected in most methylotrophs (3,10,13,15,22,42), and a few FDHs have been purified and analyzed (reviewed in reference 39). In the methylotrophic yeast Candida boidinii, the FDH step was shown not to be essential for methylotrophic growth, but FDH mutants showed reduced growth on methanol (32).…”

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confidence: 99%

Multiple Formate Dehydrogenase Enzymes in the Facultative Methylotroph Methylobacterium extorquens AM1 Are Dispensable for Growth on Methanol

Chistoserdova¹,

Laukel

Portais

et al. 2004

J Bacteriol

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“…All three enzymes differ markedly in their structure from the FDH ofMoraxella sp. (a2; Mr = 98,000 [1]) and Pseudomonas sp. strain 101 (a2; Mr = 80,000 [17]; formerly designated as methylotrophic bacterium strain 1 or Achromobacterparvulus).…”

Section: Discussionmentioning

confidence: 99%

“…Supplementation of the medium with tungstate lead to growth cessation. (2,23,26,34,50), aerobic (1,17,30,32,33,41,60) and anaerobic (3,18,36,39,59) (18,31,36,48,58).…”

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Physiological and biochemical characterization of the soluble formate dehydrogenase, a molybdoenzyme from Alcaligenes eutrophus

1993

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Organoautotrophic growth ofAlcaligenes eutrophus on formate was dependent on the presence of molybdate in the medium. Supplementation of the medium with tungstate lead to growth cessation. (2,23,26,34,50), aerobic (1,17,30,32,33,41,60) and anaerobic (3,18,36,39,59) (18,31,36,48,58).Whereas FDH from methylotrophic yeasts have very similar overall properties, considerable diversity is found for bacterial FDH concerning their size, structure, cofactor and metal content, electron acceptor spectrum, and other features (19).

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“…Among the large number of microorganisms that have formate dehydrogenase, the NAD-linked enzyme (EC 1.2.1.2) has been isolated from both facultative methylotrophic bacteria (3,13) and several yeast species (20; also reference 4 and references therein). There is a great deal of heterogeneity in the structure and composition of bacterial formate dehydrogenases.…”

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Formate dehydrogenase from the methane oxidizer Methylosinus trichosporium OB3b

1990

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Formate dehydrogenase (NAD+ depet) was isolated from the obligate methanotroph Methylosinus trichosporium OB3b. When the enzyme was isobted anaerobically, two forms of the enzyme were seen on native polyacrylamide gels, DE-52 celluloe and Sephacryl S-300 colns; they were approximately 315,000 and 155,000 daltons. The enzyme showed two subunits on sodium dodecyl sulfate-polyacrylamide gels. The Mr of the a-subunit was 53,800 ± 2,800, and that of the a-subunit was 102,600 ± 3,900. The enzyme (Mr 315,000) was composed of these subunits in an apparent a2j2 arrangemnt. Nonheme iron was present at a concentration ranging from 11 to 18 g-atoms per mol of enzyme (Mr 315,000). Among the large number of microorganisms that have formate dehydrogenase, the NAD-linked enzyme (EC 1.2.1.2) has been isolated from both facultative methylotrophic bacteria (3, 13) and several yeast species (20; also reference 4 and references therein). There is a great deal of heterogeneity in the structure and composition of bacterial formate dehydrogenases. They range from molecules with two identical subunits and no prosthetic group, found in facultative methylotrophic strains of Moraxella (30) In methylotrophs, formate dehydrogenase is a critical component involved in the oxidative metabolism of methane and methanol. The NADH generated by this enzyme is, in some cases, believed to be the cell's only source of reductant (2). In Methylosinus trichosporium, NAD-linked formate dehydrogenase functions as an in vitro electron donor to methane monooxygenase (32) and indirectly to nitrogenase via a ferredoxin-NAD+ reductase and ferredoxin (9,10) (Fig. 1). Although formate dehydrogenase plays an important role in the metabolism of nonfacultative methylotrophs (and obligate methanotrophs), this enzyme has not yet been isolated from either of these groups of bacteria. In this study we purified the formate dehydrogenase from the obligate methanotroph M. trichosporium and report on its structural and regulatory characteristics. * Corresponding author. MATERIALS AND METHODSGrowth conditions. M. trichosporium OB3b cells used for formate dehydrogenase preparations were batch cultured in 9-liter carboys on nitrate-containing medium (11) as described previously (10). In these cultures, the air-methane mixture (5:1) was dispersed by passing it through stone diffusers and stirring the culture slowly with a magnetic bar. Our cultures did not attain a high density (80 to 120 Klett units, no. 66 filter) in these batch cultures and were harvested by centrifugation after 6 days of growth. After harvesting, the cells were immediately frozen (unwashed)

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Purification and properties of formate dehydrogenase from Moraxella sp. strain C-1

Cited by 44 publications

References 28 publications

Multiple Formate Dehydrogenase Enzymes in the Facultative Methylotroph Methylobacterium extorquens AM1 Are Dispensable for Growth on Methanol

Multiple Formate Dehydrogenase Enzymes in the Facultative Methylotroph Methylobacterium extorquens AM1 Are Dispensable for Growth on Methanol

Physiological and biochemical characterization of the soluble formate dehydrogenase, a molybdoenzyme from Alcaligenes eutrophus

Formate dehydrogenase from the methane oxidizer Methylosinus trichosporium OB3b

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