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Free keywords:
Aquifex aeolicus; Complex I; Isoform enzyme; NADH:ubiquinone oxidoreductase; Bacteria
Abstract:
The NADH:ubiquinone oxidoreductase (complex I) is the first enzyme of the respiratory chain and the entry point for most electrons. Generally, the bacterial complex I consists of 14 core subunits, homologues of which are also found in complex I of mitochondria. In complex I preparations from the hyperthermophilic bacterium Aquifex aeolicus we have identified 20 partially homologous subunits by combining MALDI-TOF and LILBID mass spectrometry methods. The subunits could be assigned to two different complex I isoforms, named NQOR1 and NQOR2. NQOR1 consists of subunits NuoA2, NuoB, NuoD2, NuoE, NuoF, NuoG, NuoI1, NuoH1, NuoJ1, NuoK1, NuoL1, NuoM1 and NuoN1, with an entire mass of 504.17 kDa. NQOR2 comprises subunits NuoA1, NuoB, NuoD1, NuoE, NuoF, NuoG, NuoH2, NuoI2, NuoJ1, NuoK1, NuoL2, NuoM2 and NuoN2, with a total mass of 523.99 kDa. Three Fe-S clusters could be identified by EPR spectroscopy in a preparation containing predominantly NQOR1. These were tentatively assigned to a binuclear center N1, and two tetranuclear centers, N2 and N4. The redox midpoint potentials of N1 and N2 are −273 mV and −184 mV, respectively. Specific activity assays indicated that NQOR1 from cells grown under low concentrations of oxygen was the more active form. Increasing the concentration of oxygen in the bacterial cultures induced formation of NQOR2 showing the lower specific activity.
