Abstract
The operon of the anabolic pyruvate oxidoreductase (POR) of Methanococcus maripaludis encodes two genes (porEF) whose functions are unknown. Because these genes possess sequence similarity to polyferredoxins, they may be electron carriers to the POR. To elucidate whether the methanococcal POR requires PorEF for activity, a deletion mutant, strain JJ150, lacking porEF was constructed. Compared to the wild-type strain JJ1, the mutant grew more slowly in minimal medium and minimal plus acetate medium, and pyruvate-dependent methanogenesis was inhibited. In contrast, the methyl-viologen-dependent pyruvate-oxidation activity of POR, carbon monoxide dehydrogenase, and hydrogenase activities of the mutant were similar to those of the wild-type. Upon genetic complementation of the mutant with porEF in the methanococcal shuttle vector pMEV2+porEF, growth in minimal medium and pyruvate-dependent methanogenesis were restored to wild-type levels. Complementation with porE alone restored methanogenesis from pyruvate but not growth in minimal medium. Complementation with porF alone partially restored growth but not methanogenesis from pyruvate. Although the specific roles of porE and porF have not been determined, these results suggest that PorEF play important roles in the anabolic POR in vivo even though they are not required for the dye-dependent activity.
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Abbreviations
- CODH/ACS :
-
Carbon monoxide dehydrogenase/acetyl-CoA synthase
- POR :
-
Pyruvate oxidoreductase
References
Adams MWW, Kletzen A (1996) Oxidoreductase-type enzymes and redox proteins involved in fermentative metabolism of hyperthermophilic Archaea. Adv Protein Chem 48:101–180
Blamey JM, Adams MWW (1993) Purification and characterization of pyruvate ferredoxin oxidoreductase from the hyperthermophilic archaeon Pyrococcus furiosus. Biochim Biophys Acta 1161:19–27
Furdui C, Ragsdale SW (2000) The role of pyruvate ferredoxin oxidoreductase in pyruvate synthesis during autotrophic growth by the Wood-Ljungdahl pathway. J Biol Chem 275:28494–28499
Gardner WL (2000) Expression vectors for the methane-producing archaeon Methanococcus maripaludis. Dissertation, University of Georgia
Gardner WL, Whitman WB (1999) Expression vectors for Methanococcus maripaludis: overexpression of acetohydroxyacid synthase and Δ-galactosidase. Genetics 152:1439–1447
Gernhardt P, Possot O, Foglino M, Sibold L, Klein A (1990) Construction of an integration vector for use in the archaebacterium Methanococcus voltae and expression of a eubacterial resistance gene. Mol Gen Genet 221:273–279
Jones, WJ, Paynter MJB, Gupta R (1983) Characterization of Methanococcus maripaludis sp. nov., a new methanogen isolated from salt marsh sediment. Arch Microbiol 135:91–97
Korff, RW (1969) Purity and stability of pyruvate and Δ-ketoglutarate. Methods Enzymol 13:519–521
Lin WC, Yang YL, Whitman WB (2003) The anabolic pyruvate oxidoreductase from Methanococcus maripaludis. Arch Microbiol 179:444–456
Ragsdale SW, Ljungdahl LG (1984) Hydrogenase from Acetobacterium woodii. Arch Microbiol 139:361–365
Shieh JS, Whitman WB (1987) Pathway of acetate assimilation in autotrophic and heterotrophic methanococci. J Bacterial 167:5327–5329
Shieh J S, Whitman WB (1988) Autotrophic acetyl coenzyme A biosynthesis in Methanococcus maripaludis. J Bacteriol 170:3072–3079
Stathopoulos C, Kim WD, Li T, Anderson I, Deutsch B, Paliora S, Whitman WB, Söll D (2001) Cysteinyl-tRNA synthetase is not essential for viability of the archaeon Methanococcus maripaludis. Proc Natl Acad Sci USA 98:14292–14297
Tumbula DL, Makula RA, Whitman WB (1994) Transformation of Methanococcus maripaludis and identification of a PstI-like restriction system. FEMS Microbiol Lett 121:309–314
Tumbula DL, Keswani J, Shieh JS, Whitman WB (1995) Maintenance of methanogen stock cultures in glycerol at −70°C. In: Robb FT, Sowers KR, DasSarma S, Place AR, Schreier HJ, Fleischmann EM (eds) Archaea. A laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, pp 85–87
Whitman WB, Shieh JS, Sohn SH, Caras DS, Premachandran U (1986) Isolation and characterization of 22 mesophilic methanococci. System Appl Microbiol 7:235–240
Yang YL, Ladapo JA, Whitman WB (1992) Pyruvate oxidation by Methanococcus spp. Arch Microbiol 158:271–275
Yang YL, Glushka JN, Whitman WB (2002) Intracellular pyruvate flux in the methane-producing archaeon Methanococcus maripaludis. Arch Microbiol 178:493–498
Yoon KS, Ishii M, Kodama T, Igarashi Y (1996) Purification and characterization of pyruvate:ferredoxin oxidoreductase from Hydrogenobacter thermophilus TK-6. Arch Microbiol 167:275–279
Yoon KS, Ishii M, Kodama T, Igarashi Y (1997) Carboxylation reactions of pyruvate:ferredoxin oxidoreductase and 2-oxoglutarate:ferredoxin oxidoreductase from Hydrogenobacter thermophilus TK-6. Biosci Biotech Biochem 61:510–513
Yoon KS, Bobst C, Hemann CF, Hille R, Tabita FR (2001) Spectroscopic and functional properties of novel 2[4Fe-4S] cluster-containing ferredoxins from the green sulfur bacterium Chlorobium tepidum. J Biol Chem 276:44027–44036
Yu JP, Ladapo J, Whitman WB (1994) Pathway of glycogen metabolism in Methanococcus maripaludis. J Bacteriol 176:325–332
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This work was supported by a grant from the U.S. Department of Energy Division of Energy Biosciences.
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Lin, W., Whitman, W.B. The importance of porE and porF in the anabolic pyruvate oxidoreductase of Methanococcus maripaludis. Arch Microbiol 181, 68–73 (2004). https://doi.org/10.1007/s00203-003-0629-1
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DOI: https://doi.org/10.1007/s00203-003-0629-1