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. 1995 Aug;177(16):4748–4756. doi: 10.1128/jb.177.16.4748-4756.1995

Effects of elemental sulfur on the metabolism of the deep-sea hyperthermophilic archaeon Thermococcus strain ES-1: characterization of a sulfur-regulated, non-heme iron alcohol dehydrogenase.

K Ma 1, H Loessner 1, J Heider 1, M K Johnson 1, M W Adams 1
PMCID: PMC177241  PMID: 7642502

Abstract

The strictly anaerobic archaeon Thermococcus strain ES-1 was recently isolated from near a deep-sea hydrothermal vent. It grows at temperatures up to 91 degrees C by the fermentation of peptides and reduces elemental sulfur (S(o)) to H2S. It is shown here that the growth rates and cell yields of strain ES-1 are dependent upon the concentration of S(o) in the medium, and no growth was observed in the absence of S(o). The activities of various catabolic enzymes in cells grown under conditions of sufficient and limiting S(o) concentrations were investigated. These enzymes included alcohol dehydrogenase (ADH); formate benzyl viologen oxidoreductase; hydrogenase; glutamate dehydrogenase; alanine dehydrogenase; aldehyde ferredoxin (Fd) oxidoreductase; formaldehyde Fd oxidoreductase; and coenzyme A-dependent, Fd-linked oxidoreductases specific for pyruvate, indolepyruvate, 2-ketoglutarate, and 2-ketoisovalerate. Of these, changes were observed only with ADH, formate benzyl viologen oxidoreductase, and hydrogenase, the specific activities of which all dramatically increased in cells grown under S(o) limitation. This was accompanied by increased amounts of H2 and alcohol (ethanol and butanol) from cultures grown with limiting S(o). Such cells were used to purify ADH to electrophoretic homogeneity. ADH is a homotetramer with a subunit M(r) of 46,000 and contains 1 g-atom of Fe per subunit, which, as determined by electron paramagnetic resonance analyses, is present as a mixture of ferrous and ferric forms. No other metals or acid-labile sulfide was detected by colorimetric and elemental analyses. ADH utilized NADP(H) as a cofactor and preferentially catalyzed aldehyde reduction. It is proposed that, under So limitation, ADH reduces to alcohols the aldehydes that are generated by fermentation, thereby serving to dispose of excess reductant.

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Selected References

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  1. Adams M. W. Biochemical diversity among sulfur-dependent, hyperthermophilic microorganisms. FEMS Microbiol Rev. 1994 Oct;15(2-3):261–277. doi: 10.1111/j.1574-6976.1994.tb00139.x. [DOI] [PubMed] [Google Scholar]
  2. Adams M. W. Enzymes and proteins from organisms that grow near and above 100 degrees C. Annu Rev Microbiol. 1993;47:627–658. doi: 10.1146/annurev.mi.47.100193.003211. [DOI] [PubMed] [Google Scholar]
  3. Ammendola S., Raia C. A., Caruso C., Camardella L., D'Auria S., De Rosa M., Rossi M. Thermostable NAD(+)-dependent alcohol dehydrogenase from Sulfolobus solfataricus: gene and protein sequence determination and relationship to other alcohol dehydrogenases. Biochemistry. 1992 Dec 15;31(49):12514–12523. doi: 10.1021/bi00164a031. [DOI] [PubMed] [Google Scholar]
  4. Aono S., Bryant F. O., Adams M. W. A novel and remarkably thermostable ferredoxin from the hyperthermophilic archaebacterium Pyrococcus furiosus. J Bacteriol. 1989 Jun;171(6):3433–3439. doi: 10.1128/jb.171.6.3433-3439.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Arciero D. M., Orville A. M., Lipscomb J. D. [17O]Water and nitric oxide binding by protocatechuate 4,5-dioxygenase and catechol 2,3-dioxygenase. Evidence for binding of exogenous ligands to the active site Fe2+ of extradiol dioxygenases. J Biol Chem. 1985 Nov 15;260(26):14035–14044. [PubMed] [Google Scholar]
  6. Barrette W. C., Jr, Sawyer D. T., Fee J. A., Asada K. Potentiometric titrations and oxidation--reduction potentials of several iron superoxide dismutases. Biochemistry. 1983 Feb 1;22(3):624–627. doi: 10.1021/bi00272a015. [DOI] [PubMed] [Google Scholar]
  7. Bennetzen J. L., Hall B. D. The primary structure of the Saccharomyces cerevisiae gene for alcohol dehydrogenase. J Biol Chem. 1982 Mar 25;257(6):3018–3025. [PubMed] [Google Scholar]
  8. Blamey J. M., Adams M. W. Characterization of an ancestral type of pyruvate ferredoxin oxidoreductase from the hyperthermophilic bacterium, Thermotoga maritima. Biochemistry. 1994 Feb 1;33(4):1000–1007. doi: 10.1021/bi00170a019. [DOI] [PubMed] [Google Scholar]
  9. Blamey J. M., Adams M. W. Purification and characterization of pyruvate ferredoxin oxidoreductase from the hyperthermophilic archaeon Pyrococcus furiosus. Biochim Biophys Acta. 1993 Jan 15;1161(1):19–27. doi: 10.1016/0167-4838(93)90190-3. [DOI] [PubMed] [Google Scholar]
  10. Bleicher K., Winter J. Purification and properties of F420- and NADP(+)-dependent alcohol dehydrogenases of Methanogenium liminatans and Methanobacterium palustre, specific for secondary alcohols. Eur J Biochem. 1991 Aug 15;200(1):43–51. doi: 10.1111/j.1432-1033.1991.tb21046.x. [DOI] [PubMed] [Google Scholar]
  11. Blumentals I. I., Itoh M., Olson G. J., Kelly R. M. Role of Polysulfides in Reduction of Elemental Sulfur by the Hyperthermophilic Archaebacterium Pyrococcus furiosus. Appl Environ Microbiol. 1990 May;56(5):1255–1262. doi: 10.1128/aem.56.5.1255-1262.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Boyington J. C., Gaffney B. J., Amzel L. M. The three-dimensional structure of an arachidonic acid 15-lipoxygenase. Science. 1993 Jun 4;260(5113):1482–1486. doi: 10.1126/science.8502991. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. Bryant F. O., Adams M. W. Characterization of hydrogenase from the hyperthermophilic archaebacterium, Pyrococcus furiosus. J Biol Chem. 1989 Mar 25;264(9):5070–5079. [PubMed] [Google Scholar]
  15. Chasteen N. D., Grady J. K., Skorey K. I., Neden K. J., Riendeau D., Percival M. D. Characterization of the non-heme iron center of human 5-lipoxygenase by electron paramagnetic resonance, fluorescence, and ultraviolet-visible spectroscopy: redox cycling between ferrous and ferric states. Biochemistry. 1993 Sep 21;32(37):9763–9771. doi: 10.1021/bi00088a031. [DOI] [PubMed] [Google Scholar]
  16. Chen J. S., Mortenson L. E. Inhibition of methylene blue formation during determination of the acid-labile sulfide of iron-sulfur protein samples containing dithionite. Anal Biochem. 1977 May 1;79(1-2):157–165. doi: 10.1016/0003-2697(77)90390-6. [DOI] [PubMed] [Google Scholar]
  17. Drewke C., Ciriacy M. Overexpression, purification and properties of alcohol dehydrogenase IV from Saccharomyces cerevisiae. Biochim Biophys Acta. 1988 May 6;950(1):54–60. doi: 10.1016/0167-4781(88)90072-3. [DOI] [PubMed] [Google Scholar]
  18. Edelhoch H. Spectroscopic determination of tryptophan and tyrosine in proteins. Biochemistry. 1967 Jul;6(7):1948–1954. doi: 10.1021/bi00859a010. [DOI] [PubMed] [Google Scholar]
  19. Fischer R. J., Helms J., Dürre P. Cloning, sequencing, and molecular analysis of the sol operon of Clostridium acetobutylicum, a chromosomal locus involved in solventogenesis. J Bacteriol. 1993 Nov;175(21):6959–6969. doi: 10.1128/jb.175.21.6959-6969.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Fisher D. B., Kirkwood R., Kaufman S. Rat liver phenylalanine hydroxylase, an iron enzyme. J Biol Chem. 1972 Aug 25;247(16):5161–5167. [PubMed] [Google Scholar]
  21. Goodlove P. E., Cunningham P. R., Parker J., Clark D. P. Cloning and sequence analysis of the fermentative alcohol-dehydrogenase-encoding gene of Escherichia coli. Gene. 1989 Dec 21;85(1):209–214. doi: 10.1016/0378-1119(89)90483-6. [DOI] [PubMed] [Google Scholar]
  22. Heider J., Ma K., Adams M. W. Purification, characterization, and metabolic function of tungsten-containing aldehyde ferredoxin oxidoreductase from the hyperthermophilic and proteolytic archaeon Thermococcus strain ES-1. J Bacteriol. 1995 Aug;177(16):4757–4764. doi: 10.1128/jb.177.16.4757-4764.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hensgens C. M., Vonck J., Van Beeumen J., van Bruggen E. F., Hansen T. A. Purification and characterization of an oxygen-labile, NAD-dependent alcohol dehydrogenase from Desulfovibrio gigas. J Bacteriol. 1993 May;175(10):2859–2863. doi: 10.1128/jb.175.10.2859-2863.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kessler D., Herth W., Knappe J. Ultrastructure and pyruvate formate-lyase radical quenching property of the multienzymic AdhE protein of Escherichia coli. J Biol Chem. 1992 Sep 5;267(25):18073–18079. [PubMed] [Google Scholar]
  25. Kessler D., Leibrecht I., Knappe J. Pyruvate-formate-lyase-deactivase and acetyl-CoA reductase activities of Escherichia coli reside on a polymeric protein particle encoded by adhE. FEBS Lett. 1991 Apr 9;281(1-2):59–63. doi: 10.1016/0014-5793(91)80358-a. [DOI] [PubMed] [Google Scholar]
  26. LOVENBERG W., BUCHANAN B. B., RABINOWITZ J. C. STUDIES ON THE CHEMICAL NATURE OF CLOSTRIDIAL FERREDOXIN. J Biol Chem. 1963 Dec;238:3899–3913. [PubMed] [Google Scholar]
  27. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  28. Lamed R. J., Zeikus J. G. Novel NADP-linked alcohol--aldehyde/ketone oxidoreductase in thermophilic ethanologenic bacteria. Biochem J. 1981 Apr 1;195(1):183–190. doi: 10.1042/bj1950183. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Leonardo M. R., Cunningham P. R., Clark D. P. Anaerobic regulation of the adhE gene, encoding the fermentative alcohol dehydrogenase of Escherichia coli. J Bacteriol. 1993 Feb;175(3):870–878. doi: 10.1128/jb.175.3.870-878.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Ma K., Adams M. W. Sulfide dehydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus: a new multifunctional enzyme involved in the reduction of elemental sulfur. J Bacteriol. 1994 Nov;176(21):6509–6517. doi: 10.1128/jb.176.21.6509-6517.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Ma K., Robb F. T., Adams M. W. Purification and characterization of NADP-specific alcohol dehydrogenase and glutamate dehydrogenase from the hyperthermophilic archaeon Thermococcus litoralis. Appl Environ Microbiol. 1994 Feb;60(2):562–568. doi: 10.1128/aem.60.2.562-568.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Ma K., Schicho R. N., Kelly R. M., Adams M. W. Hydrogenase of the hyperthermophile Pyrococcus furiosus is an elemental sulfur reductase or sulfhydrogenase: evidence for a sulfur-reducing hydrogenase ancestor. Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):5341–5344. doi: 10.1073/pnas.90.11.5341. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Mai X., Adams M. W. Indolepyruvate ferredoxin oxidoreductase from the hyperthermophilic archaeon Pyrococcus furiosus. A new enzyme involved in peptide fermentation. J Biol Chem. 1994 Jun 17;269(24):16726–16732. [PubMed] [Google Scholar]
  34. Marota J. J., Shiman R. Stoichiometric reduction of phenylalanine hydroxylase by its cofactor: a requirement for enzymatic activity. Biochemistry. 1984 Mar 13;23(6):1303–1311. doi: 10.1021/bi00301a044. [DOI] [PubMed] [Google Scholar]
  35. Mukund S., Adams M. W. Characterization of a novel tungsten-containing formaldehyde ferredoxin oxidoreductase from the hyperthermophilic archaeon, Thermococcus litoralis. A role for tungsten in peptide catabolism. J Biol Chem. 1993 Jun 25;268(18):13592–13600. [PubMed] [Google Scholar]
  36. Mukund S., Adams M. W. The novel tungsten-iron-sulfur protein of the hyperthermophilic archaebacterium, Pyrococcus furiosus, is an aldehyde ferredoxin oxidoreductase. Evidence for its participation in a unique glycolytic pathway. J Biol Chem. 1991 Aug 5;266(22):14208–14216. [PubMed] [Google Scholar]
  37. Neale A. D., Scopes R. K., Kelly J. M., Wettenhall R. E. The two alcohol dehydrogenases of Zymomonas mobilis. Purification by differential dye ligand chromatography, molecular characterisation and physiological roles. Eur J Biochem. 1986 Jan 2;154(1):119–124. doi: 10.1111/j.1432-1033.1986.tb09366.x. [DOI] [PubMed] [Google Scholar]
  38. Peretz M., Burstein Y. Amino acid sequence of alcohol dehydrogenase from the thermophilic bacterium Thermoanaerobium brockii. Biochemistry. 1989 Aug 8;28(16):6549–6555. doi: 10.1021/bi00442a004. [DOI] [PubMed] [Google Scholar]
  39. Pihl T. D., Black L. K., Schulman B. A., Maier R. J. Hydrogen-oxidizing electron transport components in the hyperthermophilic archaebacterium Pyrodictium brockii. J Bacteriol. 1992 Jan;174(1):137–143. doi: 10.1128/jb.174.1.137-143.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Reid M. F., Fewson C. A. Molecular characterization of microbial alcohol dehydrogenases. Crit Rev Microbiol. 1994;20(1):13–56. doi: 10.3109/10408419409113545. [DOI] [PubMed] [Google Scholar]
  41. Rella R., Raia C. A., Pensa M., Pisani F. M., Gambacorta A., De Rosa M., Rossi M. A novel archaebacterial NAD+-dependent alcohol dehydrogenase. Purification and properties. Eur J Biochem. 1987 Sep 15;167(3):475–479. doi: 10.1111/j.1432-1033.1987.tb13361.x. [DOI] [PubMed] [Google Scholar]
  42. Riddles P. W., Blakeley R. L., Zerner B. Reassessment of Ellman's reagent. Methods Enzymol. 1983;91:49–60. doi: 10.1016/s0076-6879(83)91010-8. [DOI] [PubMed] [Google Scholar]
  43. Robb F. T., Park J. B., Adams M. W. Characterization of an extremely thermostable glutamate dehydrogenase: a key enzyme in the primary metabolism of the hyperthermophilic archaebacterium, Pyrococcus furiosus. Biochim Biophys Acta. 1992 Apr 17;1120(3):267–272. doi: 10.1016/0167-4838(92)90247-b. [DOI] [PubMed] [Google Scholar]
  44. Salerno J. C., Siedow J. N. The nature of the nitric oxide complexes of lipoxygenase. Biochim Biophys Acta. 1979 Jul 25;579(1):246–251. doi: 10.1016/0005-2795(79)90104-1. [DOI] [PubMed] [Google Scholar]
  45. Schicho R. N., Ma K., Adams M. W., Kelly R. M. Bioenergetics of sulfur reduction in the hyperthermophilic archaeon Pyrococcus furiosus. J Bacteriol. 1993 Mar;175(6):1823–1830. doi: 10.1128/jb.175.6.1823-1830.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Scopes R. K. An iron-activated alcohol dehydrogenase. FEBS Lett. 1983 Jun 13;156(2):303–306. doi: 10.1016/0014-5793(83)80517-1. [DOI] [PubMed] [Google Scholar]
  47. Stoddard B. L., Howell P. L., Ringe D., Petsko G. A. The 2.1-A resolution structure of iron superoxide dismutase from Pseudomonas ovalis. Biochemistry. 1990 Sep 25;29(38):8885–8893. doi: 10.1021/bi00490a002. [DOI] [PubMed] [Google Scholar]
  48. Thauer R. K., Jungermann K., Decker K. Energy conservation in chemotrophic anaerobic bacteria. Bacteriol Rev. 1977 Mar;41(1):100–180. doi: 10.1128/br.41.1.100-180.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Villarroya A., Juan E., Egestad B., Jörnvall H. The primary structure of alcohol dehydrogenase from Drosophila lebanonensis. Extensive variation within insect 'short-chain' alcohol dehydrogenase lacking zinc. Eur J Biochem. 1989 Mar 1;180(1):191–197. doi: 10.1111/j.1432-1033.1989.tb14632.x. [DOI] [PubMed] [Google Scholar]
  50. Wallick D. E., Bloom L. M., Gaffney B. J., Benkovic S. J. Reductive activation of phenylalanine hydroxylase and its effect on the redox state of the non-heme iron. Biochemistry. 1984 Mar 13;23(6):1295–1302. doi: 10.1021/bi00301a043. [DOI] [PubMed] [Google Scholar]
  51. Walter K. A., Bennett G. N., Papoutsakis E. T. Molecular characterization of two Clostridium acetobutylicum ATCC 824 butanol dehydrogenase isozyme genes. J Bacteriol. 1992 Nov;174(22):7149–7158. doi: 10.1128/jb.174.22.7149-7158.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Woese C. R., Kandler O., Wheelis M. L. Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4576–4579. doi: 10.1073/pnas.87.12.4576. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Wolgel S. A., Dege J. E., Perkins-Olson P. E., Jaurez-Garcia C. H., Crawford R. L., Münck E., Lipscomb J. D. Purification and characterization of protocatechuate 2,3-dioxygenase from Bacillus macerans: a new extradiol catecholic dioxygenase. J Bacteriol. 1993 Jul;175(14):4414–4426. doi: 10.1128/jb.175.14.4414-4426.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

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