Identification of a novel subunit of respiratory complex I from Thermus thermophilus.
The hydrophilic domain (peripheral arm) of the proton-translocating NADH:quinone oxidoreductase (complex I) from the thermophilic organism Thermus thermophilus HB8 has been purified and characterized. The subcomplex is stable in sodium dodecyl sulfate up to 80 degrees C. Of nine iron-sulfur clusters, four to five (one or two binuclear and three tetranuclear) could be detected by EPR in the NADH-reduced enzyme. The preparation consists of eight different polypeptides. Seven of them have been positively identified by peptide mass mapping and N-terminal sequencing as known hydrophilic subunits of T. thermophilus complex I. The eighth polypeptide copurified with the subcomplex at all stages, is strongly associated with the other subunits, and is present in crystals of the subcomplex, used for X-ray data collection. Therefore, it has been identified as a novel complex I subunit and named Nqo15. It is encoded in a locus separate from the nqo operon, containing the 14 other known complex I genes. ORFs encoding Nqo15 homologues are present in the genomes of the closest relatives of T. thermophilus. Our data show that, contrary to previous assumptions, bacterial complex I can contain proteins in addition to a "core" complement of 14 subunits.
Medical Research Council, Dunn Human Nutrition Unit, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 2XY, UK.
Philip
P
Hinchliffe
Joe
J
Carroll
Leonid A
LA
Sazanov
D000595:Amino Acid Sequence; D003460:Crystallization; D004578:Electron Spin Resonance Spectroscopy; D042967:Electron Transport Complex I / Q000737:chemistry* / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D021122:Protein Subunits / Q000737:chemistry* / Q000302:isolation & purification; D016415:Sequence Alignment; D016963:Thermus thermophilus / Q000737:chemistry*
D021122:Protein Subunits; D042967:Electron Transport Complex I