title: Oxidative-stress-inducible qorA encodes an NADPH-dependent quinone oxidoreductase catalysing a one-electron reduction in Staphylococcus aureus.
abstract: This work characterized the putative quinone oxidoreductase gene (qorA) from Staphylococcus aureus. The deduced amino acid sequence indicated that the 333 aa protein contains an NAD(P)H-binding motif. A Northern blot analysis revealed that 2.6 kb and 1.4 kb signals were detected by using a qorA probe. Both the signals were enhanced under the presence of a redox-cycling agent, 9,10-phenanthrenequinone (PQ). It was also revealed that the expression of three genes, SA1988, SA1989 (qorA) and SA1990, was enhanced at the transcriptional level by PQ exposure. The results suggested that the 2.6 kb signal detected by the qorA probe was in two co-transcripts, i.e. SA1990-qorA and qorA-SA1988 were transcribed. Besides, primer extension analyses confirmed the enhancement of qorA and SA1990 transcripts. The GST (glutathione S-transferase)-tagged QorA protein was expressed in Escherichia coli and purified using a glutathione affinity column. In purification steps, a 36 kDa band co-purified with the GST-QorA, and it was detected even in the thrombin-cleaved fraction. N-terminal amino acid sequences for the 36 kDa protein revealed that it was an intact QorA. They showed that QorA formed a multimer under physiological conditions. The purified recombinant GST-QorA catalysed NADPH consumption in the presence of PQ as a substrate, but not NADH. To characterize the catalytic activity of QorA, superoxide anion that was generated through one-electron reduction of PQ and hydroquinone that was produced by two-electron reduction of PQ were measured. During reduction of PQ by GST-QorA, superoxide anion was generated, whereas a small amount of 9,10-dihydroxyphenanthrene (hydroquinone of PQ) was produced. These results suggest that the activity of QorA is similar to zeta-Crystallin, catalysing an NADPH-dependent one-electron reduction of quinone.
authors.affiliation: Institute of Biological Sciences, University of Tsukuba, Ibaraki, 305-8577, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Maruyama
authors.affiliation: Institute of Community Medicine, University of Tsukuba, Ibaraki, 305-8577, Japan.
authors.forename: Yoshito
authors.initials: Y
authors.lastname: Kumagai
authors.affiliation: Institute of Basic Medical Sciences, University of Tsukuba, Ibaraki, 305-8577, Japan.
authors.forename: Kazuya
authors.initials: K
authors.lastname: Morikawa
authors.affiliation: Institute of Community Medicine, University of Tsukuba, Ibaraki, 305-8577, Japan.
authors.forename: Keiko
authors.initials: K
authors.lastname: Taguchi
authors.affiliation: Institute of Basic Medical Sciences, University of Tsukuba, Ibaraki, 305-8577, Japan.
authors.forename: Hideo
authors.initials: H
authors.lastname: Hayashi
authors.affiliation: Institute of Basic Medical Sciences, University of Tsukuba, Ibaraki, 305-8577, Japan.
authors.forename: Toshiko
authors.initials: T
authors.lastname: Ohta
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D016227:Benzoquinones / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D007700:Kinetics; D008969:Molecular Sequence Data; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D013211:Staphylococcus aureus / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development
chemical_list: D001426:Bacterial Proteins; D016227:Benzoquinones; C004532:quinone; D016660:NAD(P)H Dehydrogenase (Quinone)