Amino acid residues associated with cluster N3 in the NuoF subunit of the proton-translocating NADH-quinone oxidoreductase from Escherichia coli |
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Authors: | Velazquez Isabel Nakamaru-Ogiso Eiko Yano Takahiro Ohnishi Tomoko Yagi Takao |
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Institution: | Division of Biochemistry, Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA. |
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Abstract: | The NuoF subunit, which harbors NADH-binding site, of Escherichia coli NADH-quinone oxidoreductase (NDH-1) contains five conserved cysteine residues, four of which are predicted to ligate cluster N3. To determine this coordination, we overexpressed and purified the NuoF subunit and NuoF+E subcomplex in E. coli. We detected two distinct EPR spectra, arising from a 4Fe-4S] cluster (g(x,y,z)=1.90, 1.95, and 2.05) in NuoF, and a 2Fe-2S] cluster (g(x,y,z)=1.92, 1.95, and 2.01) in NuoE subunit. These clusters were assigned to clusters N3 and N1a, respectively. Based on the site-directed mutagenesis experiments, we identified that cluster N3 is ligated to the 351Cx2Cx2Cx40C398 motif. |
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Keywords: | NDH-1 bacterial proton-translocating NADH-quinone oxidoreductase EPR electron paramagnetic resonance |
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