Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1988 Jul;170(7):3072–3079. doi: 10.1128/jb.170.7.3072-3079.1988

Autotrophic acetyl coenzyme A biosynthesis in Methanococcus maripaludis.

J Shieh 1, W B Whitman 1
PMCID: PMC211251  PMID: 3133359

Abstract

To detect autotrophic CO2 assimilation in cell extracts of Methanococcus maripaludis, lactate dehydrogenase and NADH were added to convert pyruvate formed from autotrophically synthesized acetyl coenzyme A to lactate. The lactate produced was determined spectrophotometrically. When CO2 fixation was pulled in the direction of lactate synthesis, CO2 reduction to methane was inhibited. Bromoethanesulfonate (BES), a potent inhibitor of methanogenesis, enhanced lactate synthesis, and methyl coenzyme M inhibited it in the absence of BES. Lactate synthesis was dependent on CO2 and H2, but H2 + CO2-independent synthesis was also observed. In cell extracts, the rate of lactate synthesis was about 1.2 nmol min-1 mg of protein-1. When BES was added, the rate of lactate synthesis increased to 2.3 nmol min-1 mg of protein-1. Because acetyl coenzyme A did not stimulate lactate synthesis, pyruvate synthase may have been the limiting activity in these assays. Radiolabel from 14CO2 was incorporated into lactate. The percentages of radiolabel in the C-1, C-2, and C-3 positions of lactate were 73, 33, and 11%, respectively. Both carbon monoxide and formaldehyde stimulated lactate synthesis. 14CH2O was specifically incorporated into the C-3 of lactate, and 14CO was incorporated into the C-1 and C-2 positions. Low concentrations of cyanide also inhibited autotrophic growth, CO dehydrogenase activity, and autotrophic lactate synthesis. These observations are in agreement with the acetogenic pathway of autotrophic CO2 assimilation.

Full text

PDF
3072

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bonner W. M., Stedman J. D. Efficient fluorography of 3H and 14C on thin layers. Anal Biochem. 1978 Aug 15;89(1):247–256. doi: 10.1016/0003-2697(78)90747-9. [DOI] [PubMed] [Google Scholar]
  2. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  3. Daniels L., Fuchs G., Thauer R. K., Zeikus J. G. Carbon monoxide oxidation by methanogenic bacteria. J Bacteriol. 1977 Oct;132(1):118–126. doi: 10.1128/jb.132.1.118-126.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. DeMoll E., Grahame D. A., Harnly J. M., Tsai L., Stadtman T. C. Purification and properties of carbon monoxide dehydrogenase from Methanococcus vannielii. J Bacteriol. 1987 Sep;169(9):3916–3920. doi: 10.1128/jb.169.9.3916-3920.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Drake H. L., Hu S. I., Wood H. G. 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] [Google Scholar]
  6. Eikmanns B., Fuchs G., Thauer R. K. Formation of carbon monoxide from CO2 and H2 by Methanobacterium thermoautotrophicum. Eur J Biochem. 1985 Jan 2;146(1):149–154. doi: 10.1111/j.1432-1033.1985.tb08631.x. [DOI] [PubMed] [Google Scholar]
  7. Escalante-Semerena J. C., Rinehart K. L., Jr, Wolfe R. S. Tetrahydromethanopterin, a carbon carrier in methanogenesis. J Biol Chem. 1984 Aug 10;259(15):9447–9455. [PubMed] [Google Scholar]
  8. Escalante-Semerena J. C., Wolfe R. S. Tetrahydromethanopterin-dependent methanogenesis from non-physiological C1 donors in Methanobacterium thermoautotrophicum. J Bacteriol. 1985 Feb;161(2):696–701. doi: 10.1128/jb.161.2.696-701.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fuchs G., Schnitker U., Thauer R. K. Carbon monoxide oxidation by growing cultures of Clostridium pasteurianum. Eur J Biochem. 1974 Nov 1;49(1):111–115. doi: 10.1111/j.1432-1033.1974.tb03816.x. [DOI] [PubMed] [Google Scholar]
  10. Fuchs G., Stupperich E., Thauer R. K. Acetate assimilation and the synthesis of alanine, aspartate and glutamate in Methanobacterium thermoautotrophicum. Arch Microbiol. 1978 Apr 27;117(1):61–66. doi: 10.1007/BF00689352. [DOI] [PubMed] [Google Scholar]
  11. Gunsalus R. P., Wolfe R. S. Stimulation of CO2 reduction to methane by methylcoenzyme M in extracts Methanobacterium. Biochem Biophys Res Commun. 1977 Jun 6;76(3):790–795. doi: 10.1016/0006-291x(77)91570-4. [DOI] [PubMed] [Google Scholar]
  12. Hu S. I., Drake H. L., Wood H. G. 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. doi: 10.1128/jb.149.2.440-448.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hu S. I., Pezacka E., Wood H. G. Acetate synthesis from carbon monoxide by Clostridium thermoaceticum. Purification of the corrinoid protein. J Biol Chem. 1984 Jul 25;259(14):8892–8897. [PubMed] [Google Scholar]
  14. Jones W. J., Nagle D. P., Jr, Whitman W. B. Methanogens and the diversity of archaebacteria. Microbiol Rev. 1987 Mar;51(1):135–177. doi: 10.1128/mr.51.1.135-177.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kenealy W. R., Zeikus J. G. One-carbon metabolism in methanogens: evidence for synthesis of a two-carbon cellular intermediate and unification of catabolism and anabolism in Methanosarcina barkeri. J Bacteriol. 1982 Aug;151(2):932–941. doi: 10.1128/jb.151.2.932-941.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  17. Ljungdahl L. G. The autotrophic pathway of acetate synthesis in acetogenic bacteria. Annu Rev Microbiol. 1986;40:415–450. doi: 10.1146/annurev.mi.40.100186.002215. [DOI] [PubMed] [Google Scholar]
  18. Länge S., Fuchs G. Autotrophic synthesis of activated acetic acid from CO2 in Methanobacterium thermoautotrophicum. Synthesis from tetrahydromethanopterin-bound C1 units and carbon monoxide. Eur J Biochem. 1987 Feb 16;163(1):147–154. doi: 10.1111/j.1432-1033.1987.tb10748.x. [DOI] [PubMed] [Google Scholar]
  19. Ragsdale S. W., Clark J. E., Ljungdahl L. G., Lundie L. L., Drake H. L. 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] [Google Scholar]
  20. Ragsdale S. W., Wood H. G. Acetate biosynthesis by acetogenic bacteria. Evidence that carbon monoxide dehydrogenase is the condensing enzyme that catalyzes the final steps of the synthesis. J Biol Chem. 1985 Apr 10;260(7):3970–3977. [PubMed] [Google Scholar]
  21. Romesser J. A., Wolfe R. S. Coupling of methyl coenzyme M reduction with carbon dioxide activation in extracts of Methanobacterium thermoautotrophicum. J Bacteriol. 1982 Nov;152(2):840–847. doi: 10.1128/jb.152.2.840-847.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Shieh J. S., Whitman W. B. Pathway of acetate assimilation in autotrophic and heterotrophic methanococci. J Bacteriol. 1987 Nov;169(11):5327–5329. doi: 10.1128/jb.169.11.5327-5329.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Smith M. R., Lequerica J. L., Hart M. R. Inhibition of methanogenesis and carbon metabolism in Methanosarcina sp. by cyanide. J Bacteriol. 1985 Apr;162(1):67–71. doi: 10.1128/jb.162.1.67-71.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Stupperich E., Hammel K. E., Fuchs G., Thauer R. K. Carbon monoxide fixation into the carboxyl group of acetyl coenzyme A during autotrophic growth of Methanobacterium. FEBS Lett. 1983 Feb 7;152(1):21–23. doi: 10.1016/0014-5793(83)80473-6. [DOI] [PubMed] [Google Scholar]
  25. Whitman W. B., Wolfe R. S. Activation of the methylreductase system from Methanobacterium bryantii by ATP. J Bacteriol. 1983 May;154(2):640–649. doi: 10.1128/jb.154.2.640-649.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Woese C. R. Bacterial evolution. Microbiol Rev. 1987 Jun;51(2):221–271. doi: 10.1128/mr.51.2.221-271.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Zeikus J. G., Fuchs G., Kenealy W., Thauer R. K. Oxidoreductases involved in cell carbon synthesis of Methanobacterium thermoautotrophicum. J Bacteriol. 1977 Nov;132(2):604–613. doi: 10.1128/jb.132.2.604-613.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES