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The anabolic pyruvate oxidoreductase from Methanococcus maripaludis

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Abstract

In autotrophic methanogens, pyruvate oxidoreductase (POR) plays a key role in the assimilation of CO2 and the biosynthesis of organic carbon. This enzyme has been purified to homogeneity, and the genes from Methanococcus maripaludis were sequenced. The purified POR contained five polypeptides with molecular masses of 47, 33, 25, 21.5 and 13 kDa. The N-terminal sequences of four of the polypeptides had high similarity to the subunits commonly associated with this enzyme from other archaea. However, the 21.5-kDa polypeptide had not been previously observed in PORs. Nucleotide sequencing of the gene cluster encoding the POR revealed six open reading frames (porABCDEF). The genes porABCD corresponded to the subunits previously identified in PORs. On the basis of the N-terminal amino acid sequence, porE encoded the 21.5-kDa polypeptide and contained a high cysteinyl residue content and a motif indicative of a [Fe–S] cluster. porF also had a high sequence similarity to porE, a high cysteinyl residue content, and two [Fe–S] cluster motifs. Homologs to porE were also present in the genomic sequences of the autotrophic methanogens Methanocaldococcus jannaschii and Methanothermobacter thermautotrophicus. Based upon these results, it is proposed that PorE and PorF are components of a specialized system required to transfer low-potential electrons for pyruvate biosynthesis. Some biochemical properties of the purified methanococcal POR were also determined. This unstable enzyme was very sensitive to O2 and demonstrated high activity with pyruvate, oxaloacetate, and α-ketobutyrate. Methyl viologen, rubredoxin, FMN, and FAD were readily reduced. Activity was also observed with spinach and clostridial ferredoxins and cytochrome c. Coenzyme F420 was not an electron acceptor for the purified enzyme.

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Abbreviations

CODH/ACS :

Carbon monoxide dehydrogenase/acetyl-CoA synthase

PBE :

Polybuffer exchange

POR :

Pyruvate oxidoreductase

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Acknowledgements

This work was supported by a grant from the U.S. Department of Energy Division of Energy Biosciences.

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  1. Yu-Ling Yang

    Present address: Office of Science and Technology Advisors, Environmental Protection Administration, Taiwan, ROC

Authors and Affiliations

  1. Department of Microbiology, University of Georgia, Athens, GA 30602-2605, USA

    Winston C. Lin, Yu-Ling Yang & William B. Whitman

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  1. Winston C. Lin
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Correspondence to William B. Whitman.

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Lin, W.C., Yang, YL. & Whitman, W.B. The anabolic pyruvate oxidoreductase from Methanococcus maripaludis . Arch Microbiol 179, 444–456 (2003). https://doi.org/10.1007/s00203-003-0554-3

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