Production, characterization and determination of the real catalytic properties of the putative 'succinate dehydrogenase' from Wolinella succinogenes.
Both the genomes of the epsilonproteobacteria Wolinella succinogenes and Campylobacter jejuni contain operons (sdhABE) that encode for so far uncharacterized enzyme complexes annotated as 'non-classical' succinate:quinone reductases (SQRs). However, the role of such an enzyme ostensibly involved in aerobic respiration in an anaerobic organism such as W. succinogenes has hitherto been unknown. We have established the first genetic system for the manipulation and production of a member of the non-classical succinate:quinone oxidoreductase family. Biochemical characterization of the W. succinogenes enzyme reveals that the putative SQR is in fact a novel methylmenaquinol:fumarate reductase (MFR) with no detectable succinate oxidation activity, clearly indicative of its involvement in anaerobic metabolism. We demonstrate that the hydrophilic subunits of the MFR complex are, in contrast to all other previously characterized members of the superfamily, exported into the periplasm via the twin-arginine translocation (tat)-pathway. Furthermore we show that a single amino acid exchange (Ala86-->His) in the flavoprotein of that enzyme complex is the only additional requirement for the covalent binding of the otherwise non-covalently bound FAD. Our results provide an explanation for the previously published puzzling observation that the C. jejuni sdhABE operon is upregulated in an oxygen-limited environment as compared with microaerophilic laboratory conditions.
Juhnke
Hanno D
HD
Cluster of Excellence 'Macromolecular Complexes', Max Planck Institute of Biophysics, Department of Molecular Membrane Biology, Max-von-Laue-Str. 3, D-60438 Frankfurt am Main, Germany.
Hiltscher
Heiko
H
Nasiri
Hamid R
HR
Schwalbe
Harald
H
Lancaster
C Roy D
CR
D001692:Biological Transport; D002384:Catalysis; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009876:Operon; D019897:Periplasm / Q000378:metabolism; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D011809:Quinones / Q000378:metabolism; D013385:Succinate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D016968:Wolinella / Q000201:enzymology* / Q000235:genetics
D021122:Protein Subunits; D011809:Quinones; D013385:Succinate Dehydrogenase