The proton-translocating NADH-quinone oxidoreductase (NDH-1) of thermophilic bacterium Thermus thermophilus HB-8. Complete DNA sequence of the gene cluster and thermostable properties of the expressed NQO2 subunit.
The genes encoding the proton-translocating NADH-quinone oxidoreductase (NDH-1) of a thermophilic bacterium Thermus thermophilus HB-8 were cloned and sequenced. They constitute a cluster that is composed of 14 structural genes and contains no unidentified reading frames. All of the 14 structural genes, which are designated NQO1-14, encode subunits homologous to those of Paracoccus denitrificans NDH-1, respectively, and are arranged in the same order as other bacterial NDH-1 genes. T. thermophilus NDH-1 contains at most nine putative iron-sulfur cluster binding sites, eight of which are commonly found in other organisms. The T. thermophilus NQO2 subunit was expressed in Escherichia coli. The expressed subunit bears a single [2Fe-2S] cluster whose optical and EPR properties are very similar to those of N1a cluster in the P. denitrificans NQO2 subunit (Yano, T., Sled', V.D., Ohnishi, T., and Yagi, T. (1994) Biochemistry 33, 494-499). These results strongly suggest that the T. thermophilus NDH-1 is similar to other NDH-1 enzyme complexes in terms of subunit and cofactor composition. The T. thermophilus NQO2 subunit displayed much higher stability than the mesophilic equivalent and its iron-sulfur cluster remained intact even after incubation for 3 h at 65 degrees C under anaerobic conditions. With the advantage of thermostability, the T. thermophilus NDH-1 provides a great model system to investigate the structure-function relationship of the NDH-1 enzyme complexes.
Yano
T
T
Division of Biochemistry, Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, California 92037, USA.
Chu
S S
SS
Sled'
V D
VD
Ohnishi
T
T
Yagi
T
T
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001692:Biological Transport; D002417:Cattle; D004795:Enzyme Stability; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D005810:Multigene Family; D011522:Protons; D011808:Quinone Reductases / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D016963:Thermus thermophilus / Q000201:enzymology*
D011522:Protons; C041831:NADH dehydrogenase (quinone); D011808:Quinone Reductases