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J Bacteriol. Sep 1983; 155(3): 1224–1237.
PMCID: PMC217820

Isolation of carbon monoxide dehydrogenase from Acetobacterium woodii and comparison of its properties with those of the Clostridium thermoaceticum enzyme.

Abstract

An oxygen-labile carbon monoxide dehydrogenase was purified to at least 98% homogeneity from fructose-grown cells of Acetobacterium woodii. Gel filtration and electrophoresis experiments gave molecular weights of 480,000 and 153,000, respectively, of the active enzyme. The molecular weights for the subunits are 80,000 and 68,000; the subunits occur in equal proportion. The small subunit of the A. woodii enzyme differs in size from that of the Clostridium thermoaceticum enzyme; however, the large subunits are similar. The specific activity of the A. woodii enzyme, measured at 30 degrees C and pH 7.6, is 500 mumol of CO oxidized min-1 mg-1 with 20 mM methyl viologen as the electron acceptor. Analysis revealed (number per dimer) iron (9), acid-labile sulfide (12), nickel (1.4), and magnesium or zinc (1). This metal content is quite similar to that of the C. thermoaceticum enzyme (Ragsdale et al., J. Biol. Chem. 258:2364-2369, 1983). The nickel as well as the iron-sulfur clusters are redox-active, as was found for the C. thermoaceticum enzyme (Ragsdale et al., Biochem. Biophys. Res. Commun. 108:658-663, 1982). CO can reduce and CO2 can oxidize the iron-sulfur clusters. The enzyme is inhibited by cyanide, but CO2 in the presence of reduced methyl viologen or CO alone can reverse or prevent this inhibition. Several ferredoxins, flavodoxin, and rubredoxin and some artificial electron carriers were tested for their relative rates of reaction with the CO dehydrogenases from A. woodii, C. thermoaceticum, and Clostridium formicoaceticum. Rubredoxin was by far the most reactive acceptor and is proposed to be the primary natural electron carrier for the acetogenic CO dehydrogenases.

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  • Andreesen JR, Gottschalk G, Schlegel HG. Clostridium formicoaceticum nov. spec. isolation, description and distinction from C. aceticum and C. thermoaceticum. Arch Mikrobiol. 1970;72(2):154–174. [PubMed]
  • Andrews P. Estimation of the molecular weights of proteins by Sephadex gel-filtration. Biochem J. 1964 May;91(2):222–233. [PMC free article] [PubMed]
  • Babior BM, Moss TH, Orme-Johnson WH, Beinert H. The mechanism of action of ethanolamine ammonia-lyase, a B-12-dependent enzyme. The participation of paramagnetic species in the catalytic deamination of 2-aminopropanol. J Biol Chem. 1974 Jul 25;249(14):4537–4544. [PubMed]
  • Chrambach A. Electrophoresis and electrofocusing on polyacrylamide gel in the study of native macromolecules. Mol Cell Biochem. 1980 Jan 16;29(1):23–46. [PubMed]
  • Clark JE, Ljungdahl LG. Purification and properties of 5,10-methenyltetrahydrofolate cyclohydrolase from Clostridium formicoaceticum. J Biol Chem. 1982 Apr 10;257(7):3833–3836. [PubMed]
  • Clark JE, Ragsdale SW, Ljungdahl LG, Wiegel J. Levels of enzymes involved in the synthesis of acetate from CO2 in Clostridium thermoautotrophicum. J Bacteriol. 1982 Jul;151(1):507–509. [PMC free article] [PubMed]
  • DAVIS BJ. DISC ELECTROPHORESIS. II. METHOD AND APPLICATION TO HUMAN SERUM PROTEINS. Ann N Y Acad Sci. 1964 Dec 28;121:404–427. [PubMed]
  • Diekert G, Ritter M. Nickel requirement of Acetobacterium woodii. J Bacteriol. 1982 Aug;151(2):1043–1045. [PMC free article] [PubMed]
  • Diekert GB, Thauer RK. Carbon monoxide oxidation by Clostridium thermoaceticum and Clostridium formicoaceticum. J Bacteriol. 1978 Nov;136(2):597–606. [PMC free article] [PubMed]
  • DOEG KA, ZIEGLER DM. Simplified methods for the estimation of iron in mitochondria and submitochondrial fractions. Arch Biochem Biophys. 1962 Apr;97:37–40. [PubMed]
  • Drake HL. Occurrence of nickel in carbon monoxide dehydrogenase from Clostridium pasteurianum and Clostridium thermoaceticum. J Bacteriol. 1982 Feb;149(2):561–566. [PMC free article] [PubMed]
  • Drake HL, Hu SI, Wood HG. Purification of carbon monoxide dehydrogenase, a nickel enzyme from Clostridium thermocaceticum. J Biol Chem. 1980 Aug 10;255(15):7174–7180. [PubMed]
  • Drake HL, Hu SI, Wood HG. Purification of five components from Clostridium thermoaceticum which catalyze synthesis of acetate from pyruvate and methyltetrahydrofolate. Properties of phosphotransacetylase. J Biol Chem. 1981 Nov 10;256(21):11137–11144. [PubMed]
  • Elliott JI, Brewer JM. The inactivation of yeast enolase by 2,3-butanedione. Arch Biochem Biophys. 1978 Sep;190(1):351–357. [PubMed]
  • Elliott JI, Ljungdahl LG. Isolation and characterization of an Fe,-S8 ferredoxin (ferredoxin II) from Clostridium thermoaceticum. J Bacteriol. 1982 Jul;151(1):328–333. [PMC free article] [PubMed]
  • Fernández VM, Gutiérrez C, Ballesteros A. Determination of hydrogenase activity using an anaerobic spectrophotometric device. Anal Biochem. 1982 Feb;120(1):85–90. [PubMed]
  • Fuchs G, Schnitker U, Thauer RK. Carbon monoxide oxidation by growing cultures of Clostridium pasteurianum. Eur J Biochem. 1974 Nov 1;49(1):111–115. [PubMed]
  • Hedrick JL, Smith AJ. Size and charge isomer separation and estimation of molecular weights of proteins by disc gel electrophoresis. Arch Biochem Biophys. 1968 Jul;126(1):155–164. [PubMed]
  • Hu SI, Drake HL, Wood HG. Synthesis of acetyl coenzyme A from carbon monoxide, methyltetrahydrofolate, and coenzyme A by enzymes from Clostridium thermoaceticum. J Bacteriol. 1982 Feb;149(2):440–448. [PMC free article] [PubMed]
  • Jungermann K, Thauer RK, Leimenstoll G, Decker K. Function of reduced pyridine nucleotide-ferredoxin oxidoreductases in saccharolytic Clostridia. Biochim Biophys Acta. 1973 May 30;305(2):268–280. [PubMed]
  • Kim YM, Hegeman GD. Purification and some properties of carbon monoxide dehydrogenase from Pseudomonas carboxydohydrogena. J Bacteriol. 1981 Dec;148(3):904–911. [PMC free article] [PubMed]
  • Kim YM, Hegeman GD. Electron transport system of an aerobic carbon monoxide-oxidizing bacterium. J Bacteriol. 1981 Dec;148(3):991–994. [PMC free article] [PubMed]
  • Kojima N, Fox JA, Hausinger RP, Daniels L, Orme-Johnson WH, Walsh C. Paramagnetic centers in the nickel-containing, deazaflavin-reducing hydrogenase from Methanobacterium thermoautotrophicum. Proc Natl Acad Sci U S A. 1983 Jan;80(2):378–382. [PMC free article] [PubMed]
  • Krüger HJ, Huynh BH, Ljungdahl PO, Xavier AV, Der Vartanian DV, Moura I, Peck HD, Jr, Teixeira M, Moura JJ, LeGall J. Evidence for nickel and a three-iron center in the hydrogenase of Desulfovibrio desulfuricans. J Biol Chem. 1982 Dec 25;257(24):14620–14623. [PubMed]
  • Gall JL, Forget N. Purification of electron-transfer components from sulfate-reducing bacteria. Methods Enzymol. 1978;53:613–634. [PubMed]
  • Ljungdahl LG, Andreesen JR. Formate dehydrogenase, a selenium--tungsten enzyme from Clostridium thermoaceticum. Methods Enzymol. 1978;53:360–372. [PubMed]
  • Lovenberg W, Sobel BE. Rubredoxin: a new electron transfer protein from Clostridium pasteurianum. Proc Natl Acad Sci U S A. 1965 Jul;54(1):193–199. [PMC free article] [PubMed]
  • Lynd L, Kerby R, Zeikus JG. Carbon monoxide metabolism of the methylotrophic acidogen Butyribacterium methylotrophicum. J Bacteriol. 1982 Jan;149(1):255–263. [PMC free article] [PubMed]
  • Meyer O. Chemical and spectral properties of carbon monoxide: methylene blue oxidoreductase. The molybdenum-containing iron-sulfur flavoprotein from Pseudomonas carboxydovorans. J Biol Chem. 1982 Feb 10;257(3):1333–1341. [PubMed]
  • Meyer O, Schlegel HG. Oxidation of carbon monoxide in cell extracts of Pseudomonas carboxydovorans. J Bacteriol. 1979 Feb;137(2):811–817. [PMC free article] [PubMed]
  • Meyer O, Schlegel HG. Carbon monoxide:methylene blue oxidoreductase from Pseudomonas carboxydovorans. J Bacteriol. 1980 Jan;141(1):74–80. [PMC free article] [PubMed]
  • Moore MR, O'Brien WE, Ljungdahl LG. Purification and characterization of nicotinamide adenine dinucleotide-dependent methylenetetrahydrofolate dehydrogenase from Clostridium formicoaceticum. J Biol Chem. 1974 Aug 25;249(16):5250–5253. [PubMed]
  • Moura JJ, Moura I, Huynh BH, Krüger HJ, Teixeira M, DuVarney RC, DerVartanian DV, Xavier AV, Peck HD, Jr, LeGall J. Unambiguous identification of the nickel EPR signal in 61Ni-enriched Desulfovibrio gigas hydrogenase. Biochem Biophys Res Commun. 1982 Oct 29;108(4):1388–1393. [PubMed]
  • Petitdemange H, Marczak R, Blusson H, Gay R. Isolation and properties of reduced nicotinamide adenine dinucleotiderubredoxin oxidoreductase of Clostridium acetobutylicum. Biochem Biophys Res Commun. 1979 Dec 28;91(4):1258–1265. [PubMed]
  • Rabinowitz JC. Analysis of acid-labile sulfide and sulfhydryl groups. Methods Enzymol. 1978;53:275–277. [PubMed]
  • Ragsdale SW, Clark JE, Ljungdahl LG, Lundie LL, Drake HL. Properties of purified carbon monoxide dehydrogenase from Clostridium thermoaceticum, a nickel, iron-sulfur protein. J Biol Chem. 1983 Feb 25;258(4):2364–2369. [PubMed]
  • Reisner AH, Nemes P, Bucholtz C. The use of Coomassie Brilliant Blue G250 perchloric acid solution for staining in electrophoresis and isoelectric focusing on polyacrylamide gels. Anal Biochem. 1975 Apr;64(2):509–516. [PubMed]
  • Uffen RL. Anaerobic growth of a Rhodopseudomonas species in the dark with carbon monoxide as sole carbon and energy substrate. Proc Natl Acad Sci U S A. 1976 Sep;73(9):3298–3302. [PMC free article] [PubMed]
  • Weber K, Pringle JR, Osborn M. Measurement of molecular weights by electrophoresis on SDS-acrylamide gel. Methods Enzymol. 1972;26:3–27. [PubMed]
  • Yamamoto I, Saiki T, Liu SM, Ljungdahl LG. Purification and properties of NADP-dependent formate dehydrogenase from Clostridium thermoaceticum, a tungsten-selenium-iron protein. J Biol Chem. 1983 Feb 10;258(3):1826–1832. [PubMed]
  • Yang SS, Ljungdahl LG, LeGall J. A four-iron, four-sulfide ferredoxin with high thermostability from Clostridium thermoaceticum. J Bacteriol. 1977 Jun;130(3):1084–1090. [PMC free article] [PubMed]

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