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Oxaloacetate Synthesis in the Methanarchaeon Methanosarcina barkeri: Pyruvate Carboxylase Genes and a Putative Escherichia coli-Type Bifunctional Biotin Protein Ligase Gene (bpl/birA) Exhibit a Unique Organization

American Society for Microbiology
Publication Date
  • Enzymes And Proteins
  • Archaeology
  • Biology


Evidence is presented that, in Methanosarcina barkeri oxaloacetate synthesis, an essential and major CO2 fixation reaction is catalyzed by an apparent α4β4-type acetyl coenzyme A-independent pyruvate carboxylase (PYC), composed of 64.2-kDa biotinylated and 52.9-kDa ATP-binding subunits. The purified enzyme was most active at 70°C, insensitive to aspartate and glutamate, mildly inhibited by α-ketoglutarate, and severely inhibited by ATP, ADP, and excess Mg2+. It showed negative cooperativity towards bicarbonate at 70°C but not at 37°C. The organism expressed holo-PYC without an external supply of biotin and, thus, synthesized biotin. pycA, pycB, and a putative bpl gene formed a novel operon-like arrangement. Unlike other archaeal homologs, the putative biotin protein ligases (BPLs) of M. barkeri and the closely related euryarchaeon Archaeoglobus fulgidus appeared to be of the Escherichia coli-type (bifunctional, with two activities: BirA or a repressor of the biotin operon and BPL). We found the element Tyr(Phe)ProX5Phe(Tyr) to be fully conserved in biotin-dependent enzymes; it might function as the hinge for their “swinging arms.”

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