title: Crystal structures of the quinone oxidoreductase from Thermus thermophilus HB8 and its complex with NADPH: implication for NADPH and substrate recognition.
abstract: The crystal structures of the zeta-crystalline-like soluble quinone oxidoreductase from Thermus thermophilus HB8 (QOR(Tt)) and of its complex with NADPH have been determined at 2.3- and 2.8-A resolutions, respectively. QOR(Tt) is composed of two domains, and its overall fold is similar to the folds of Escherichia coli quinone oxidoreductase (QOR(Ec)) and horse liver alcohol dehydrogenase. QOR(Tt) forms a homodimer in the crystal by interaction of the betaF-strands in domain II, forming a large beta-sheet that crosses the dimer interface. High thermostability of QOR(Tt) was evidenced by circular dichroic measurement. NADPH is located between the two domains in the QOR(Tt)-NADPH complex. The disordered segment involved in the coenzyme binding of apo-QOR(Tt) becomes ordered upon NADPH binding. The segment covers an NADPH-binding cleft and may serve as a lid. The 2'-phosphate group of the adenine of NADPH is surrounded by polar and positively charged residues in QOR(Tt), suggesting that QOR(Tt) binds NADPH more readily than NADH. The putative substrate-binding site of QOR(Tt), unlike that of QOR(Ec), is largely blocked by nearby residues, permitting access only to small substrates. This may explain why QOR(Tt) has weak p-benzoquinone reduction activity and is inactive with such large substrates of QOR(Ec) as 5-hydroxy-1,4-naphthoquinone and phenanthraquinone.
authors.affiliation: Department of Biology, Graduate School of Science, Osaka University, Toyonaka, Osaka 560-0043, Japan.
authors.forename: Yoshimitsu
authors.initials: Y
authors.lastname: Shimomura
authors.forename: Yoshimitsu
authors.initials: Y
authors.lastname: Kakuta
authors.forename: Keiichi
authors.initials: K
authors.lastname: Fukuyama
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002942:Circular Dichroism; D003460:Crystallization; D018360:Crystallography, X-Ray; D019281:Dimerization; D004795:Enzyme Stability; D006860:Hydrogen Bonding; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000737:chemistry* / Q000378:metabolism; D009249:NADP / Q000378:metabolism*; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D013379:Substrate Specificity; D013696:Temperature; D016963:Thermus thermophilus / Q000201:enzymology*
chemical_list: D009249:NADP; D016660:NAD(P)H Dehydrogenase (Quinone)