title: Decarboxylating and nondecarboxylating glutaryl-coenzyme A dehydrogenases in the aromatic metabolism of obligately anaerobic bacteria.
abstract: In anaerobic bacteria using aromatic growth substrates, glutaryl-coenzyme A (CoA) dehydrogenases (GDHs) are involved in the catabolism of the central intermediate benzoyl-CoA to three acetyl-CoAs and CO(2). In this work, we studied GDHs from the strictly anaerobic, aromatic compound-degrading organisms Geobacter metallireducens (GDH(Geo)) (Fe[III] reducing) and Desulfococcus multivorans (GDH(Des)) (sulfate reducing). GDH(Geo) was purified from cells grown on benzoate and after the heterologous expression of the benzoate-induced bamM gene. The gene coding for GDH(Des) was identified after screening of a cosmid gene library. Reverse transcription-PCR revealed that its expression was induced by benzoate; the product was heterologously expressed and isolated. Both wild-type and recombinant GDH(Geo) catalyzed the oxidative decarboxylation of glutaryl-CoA to crotonyl-CoA at similar rates. In contrast, recombinant GDH(Des) catalyzed only the dehydrogenation to glutaconyl-CoA. The latter compound was decarboxylated subsequently to crotonyl-CoA by the addition of membrane extracts from cells grown on benzoate in the presence of 20 mM NaCl. All GDH enzymes were purified as homotetramers of a 43- to 44-kDa subunit and contained 0.6 to 0.7 flavin adenine dinucleotides (FADs)/monomer. The kinetic properties for glutaryl-CoA conversion were as follows: for GDH(Geo), the K(m) was 30 +/- 2 microM and the V(max) was 3.2 +/- 0.2 micromol min(-1) mg(-1), and for GDH(Des), the K(m) was 52 +/- 5 microM and the V(max) was 11 +/- 1 micromol min(-1) mg(-1). GDH(Des) but not GDH(Geo) was inhibited by glutaconyl-CoA. Highly conserved amino acid residues that were proposed to be specifically involved in the decarboxylation of the intermediate glutaconyl-CoA were identified in GDH(Geo) but are missing in GDH(Des). The differential use of energy-yielding/energy-demanding enzymatic processes in anaerobic bacteria that degrade aromatic compounds is discussed in view of phylogenetic relationships and constraints of overall energy metabolism.
authors.affiliation: Institute of Biochemistry, University of Leipzig, Leipzig, Germany.
authors.forename: Simon
authors.initials: S
authors.lastname: Wischgoll
authors.forename: Martin
authors.initials: M
authors.lastname: Taubert
authors.forename: Franziska
authors.initials: F
authors.lastname: Peters
authors.forename: Nico
authors.initials: N
authors.lastname: Jehmlich
authors.forename: Martin
authors.initials: M
authors.lastname: von Bergen
authors.forename: Matthias
authors.initials: M
authors.lastname: Boll
mesh_terms: D000214:Acyl Coenzyme A / Q000737:chemistry / Q000378:metabolism; D000693:Anaerobiosis / Q000235:genetics; D001565:Benzoates / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D020564:Deltaproteobacteria / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects / Q000235:genetics; D045302:Geobacter / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D050770:Glutaryl-CoA Dehydrogenase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000214:Acyl Coenzyme A; D001565:Benzoates; C032423:glutaconyl-coenzyme A; C037817:benzoyl-coenzyme A; D050770:Glutaryl-CoA Dehydrogenase