title: Enzymology and evolution of the pyruvate pathway to 2-oxobutyrate in Methanocaldococcus jannaschii.
abstract: The archaeon Methanocaldococcus jannaschii uses three different 2-oxoacid elongation pathways, which extend the chain length of precursors in leucine, isoleucine, and coenzyme B biosyntheses. In each of these pathways an aconitase-type hydrolyase catalyzes an hydroxyacid isomerization reaction. The genome sequence of M. jannaschii encodes two homologs of each large and small subunit that forms the hydrolyase, but the genes are not cotranscribed. The genes are more similar to each other than to previously characterized isopropylmalate isomerase or homoaconitase enzyme genes. To identify the functions of these homologs, the four combinations of subunits were heterologously expressed in Escherichia coli, purified, and reconstituted to generate the iron-sulfur center of the holoenzyme. Only the combination of MJ0499 and MJ1277 proteins catalyzed isopropylmalate and citramalate isomerization reactions. This pair also catalyzed hydration half-reactions using citraconate and maleate. Another broad-specificity enzyme, isopropylmalate dehydrogenase (MJ0720), catalyzed the oxidative decarboxylation of beta-isopropylmalate, beta-methylmalate, and d-malate. Combined with these results, phylogenetic analysis suggests that the pyruvate pathway to 2-oxobutyrate (an alternative to threonine dehydratase in isoleucine biosynthesis) evolved several times in bacteria and archaea. The enzymes in the isopropylmalate pathway of leucine biosynthesis facilitated the evolution of 2-oxobutyrate biosynthesis through the introduction of a citramalate synthase, either by gene recruitment or gene duplication and functional divergence.
authors: Department of Chemistry and Biochemistry, University of Texas at Austin, 1 University Station A5300, Austin, TX 78712, USA.
authors: Randy M
authors: RM
authors: Drevland
authors: Abdul
authors: A
authors: Waheed
authors: David E
authors: DE
authors: Graham
mesh_terms: D050539:3-Isopropylmalate Dehydrogenase / Q000378:metabolism; D000154:Aconitate Hydratase / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000235:genetics / Q000378:metabolism*; D002087:Butyrates / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D019143:Evolution, Molecular; D006836:Hydro-Lyases / Q000378:metabolism; D007535:Isomerases / Q000378:metabolism; D008293:Malates / Q000378:metabolism; D008298:Maleates / Q000378:metabolism; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D017015:Methanococcales / Q000145:classification / Q000201:enzymology* / Q000235:genetics*; D008969:Molecular Sequence Data; D010802:Phylogeny; D021122:Protein Subunits; D019289:Pyruvic Acid / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D019843:Archaeal Proteins; D002087:Butyrates; D008293:Malates; D008298:Maleates; D021122:Protein Subunits; D011994:Recombinant Proteins; C005087:alpha-ketobutyric acid; D019289:Pyruvic Acid; D050539:3-Isopropylmalate Dehydrogenase; D006836:Hydro-Lyases; D000154:Aconitate Hydratase; C021529:maleate hydratase; C032241:isopropylmalate isomerase; C019905:(R)-2-methylmalate dehydratase; D007535:Isomerases