title: Genes, enzymes, and regulation of para-cresol metabolism in Geobacter metallireducens.
abstract: In aerobic and facultatively anaerobic bacteria, the degradation of para-cresol (p-cresol) involves the initial hydroxylation to p-hydroxybenzyl alcohol by water catalyzed by the soluble, periplasmatic flavocytochrome p-cresol methylhydroxylase (PCMH; alpha(2)beta(2) composition). In denitrifying bacteria the further metabolism proceeds via oxidation to p-hydroxybenzoate, the formation of p-hydroxybenzoyl-coenzyme A (CoA), and the subsequent dehydroxylation of the latter to benzoyl-CoA by reduction. In contrast, the strictly anaerobic Desulfobacterium cetonicum degrades p-cresol by addition to fumarate, yielding p-hydroxybenzylsuccinate. In this work, in vitro enzyme activity measurements revealed that the obligately anaerobic Geobacter metallireducens uses the p-cresol degradation pathway of denitrifying bacteria. Surprisingly, PCMH, which is supposed to catalyze both p-cresol hydroxylation and p-hydroxybenzyl alcohol oxidation to the corresponding aldehyde, was located in the membrane fraction. The alpha subunit of the enzyme was present in two isoforms, suggesting an alphaalpha'beta(2) composition. We propose that the unusual asymmetric architecture and the membrane association of PCMH might be important for alternative electron transfer routes to either cytochrome c (in the case of p-cresol oxidation) or to menaquinone (in the case of p-hydroxybenzyl alcohol oxidation). Unusual properties of further enzymes of p-cresol metabolism, p-hydroxybenzoate-CoA ligase, and p-hydroxybenzoyl-CoA reductase were identified and are discussed. A proteomic approach identified a gene cluster comprising most of the putative structural genes for enzymes involved in p-cresol metabolism (pcm genes). Reverse transcription-PCR studies revealed a different regulation of transcription of pcm genes and the corresponding enzyme activities, suggesting the presence of posttranscriptional regulatory elements.
authors: Institute for Biochemistry, Brüderstrasse 34, D-04103 Leipzig, Germany.
authors: Franziska
authors: F
authors: Peters
authors: Dimitri
authors: D
authors: Heintz
authors: Jörg
authors: J
authors: Johannes
authors: Alain
authors: A
authors: van Dorsselaer
authors: Matthias
authors: M
authors: Boll
mesh_terms: D000214:Acyl Coenzyme A / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003408:Cresols / Q000737:chemistry / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D015964:Gene Expression Regulation, Bacterial; D023061:Gene Order; D005798:Genes, Bacterial; D045302:Geobacter / Q000235:genetics / Q000378:metabolism*; D062385:Hydroxybenzoates / Q000737:chemistry / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000378:metabolism; D015394:Molecular Structure; D005810:Multigene Family; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D000214:Acyl Coenzyme A; D001426:Bacterial Proteins; D003408:Cresols; D062385:Hydroxybenzoates; D007527:Isoenzymes; C032538:4-cresol; C037817:benzoyl-coenzyme A; D006899:Mixed Function Oxygenases; D010088:Oxidoreductases; C049502:4-cresol dehydrogenase (hydroxylating)