Publications

• Molecular cloning of the isoquinoline 1-oxidoreductase genes from Pseudomonas diminuta 7, structural analysis of iorA and iorB, and sequence comparisons with other molybdenum-containing hydroxylases.

  Lehmann M., Tshisuaka B., Fetzner S., Lingens F.Y.

  The iorA and iorB genes from the isoquinoline-degrading bacterium Pseudomonas diminuta 7, encoding the heterodimeric molybdo-iron-sulfur-protein isoquinoline 1-oxidoreductase, were cloned and sequenced. The deduced amino acid sequences IorA and IorB showed homologies (i) to the small (gamma) and l ... >> More

  The iorA and iorB genes from the isoquinoline-degrading bacterium Pseudomonas diminuta 7, encoding the heterodimeric molybdo-iron-sulfur-protein isoquinoline 1-oxidoreductase, were cloned and sequenced. The deduced amino acid sequences IorA and IorB showed homologies (i) to the small (gamma) and large (alpha) subunits of complex molybdenum-containing hydroxylases (alpha beta gamma/alpha 2 beta 2 gamma 2) possessing a pterin molybdenum cofactor with a monooxo-monosulfido-type molybdenum center, (ii) to the N- and C-terminal regions of aldehyde oxidoreductase from Desulfovibrio gigas, and (iii) to the N- and C-terminal domains of eucaryotic xanthine dehydrogenases, respectively. The closest similarity to IorB was shown by aldehyde dehydrogenase (Adh) from the acetic acid bacterium Acetobacter polyoxogenes. Five conserved domains of IorB were identified by multiple sequence alignments. Whereas IorB and Adh showed an identical sequential arrangement of these conserved domains, in all other molybdenum-containing hydroxylases the relative position of "domain A" differed. IorA contained eight conserved cysteine residues. The amino acid pattern harboring the four cysteine residues proposed to ligate the Fe/S I cluster was homologous to the consensus binding site of bacterial and chloroplast-type [2Fe-2S] ferredoxins, whereas the pattern including the four cysteines assumed to ligate the Fe/S II center showed no similarities to any described [2Fe-2S] binding motif. The N-terminal region of IorB comprised a putative signal peptide similar to typical leader peptides, indicating that isoquinoline 1-oxidoreductase is associated with the cell membrane. << Less

  J. Biol. Chem. 270:14420-14429(1995) [PubMed] [EuropePMC]

• Purification and characterization of isoquinoline 1-oxidoreductase from Pseudomonas diminuta 7, a novel molybdenum-containing hydroxylase.

  Lehmann M., Tshisuaka B., Fetzner S., Roger P., Lingens F.Y.

  Isoquinoline 1-oxidoreductase, which catalyzes the hydroxylation of isoquinoline to 1-oxo-1,2-dihydroisoquinoline with concomitant reduction of a suitable electron acceptor, was purified from the isoquinoline degrading bacterium Pseudomonas diminuta 7 to apparent homogeneity. The native enzyme was ... >> More

  Isoquinoline 1-oxidoreductase, which catalyzes the hydroxylation of isoquinoline to 1-oxo-1,2-dihydroisoquinoline with concomitant reduction of a suitable electron acceptor, was purified from the isoquinoline degrading bacterium Pseudomonas diminuta 7 to apparent homogeneity. The native enzyme was a heterodimer with a molecular mass of 95 kDa consisting of a 16- and a 80-kDa subunit. It contained 0.85 g atom molybdenum, 3.95 g atom iron, 3.9 g atom acid-labile sulfur, 2.1 mol of phosphate, and 1 mol of CMP/mol of enzyme. CMP and phosphate are suggested to originate from molybdopterin cytosine dinucleotide of the pterin molybdenum cofactor. It is assumed that the iron and the acid-labile sulfur are arranged in two (2Fe-2S) clusters. The isoelectric point of the isoquinoline 1-oxidoreductase was within the range of pH 6.2 to 6.8. Cytochrome c, ferricyanide, and several non-physiological electron acceptors served as oxidizing substrates, whereas O2 and NAD were not used. Isoquinoline 1-oxidoreductase revealed a high specificity toward the reducing substrates isoquinoline, 5-hydroxyisoquinoline, quinazoline, and phthalazine. Isoquinoline 1-oxidoreductase was inactivated by methanol, arsenite, p-hydroxymercuribenzoate, 1,10-phenanthroline, and cyanide. Additionally, the enzyme was inactivated upon incubation with its substrates isoquinoline, which slowly inhibited the enzyme in the absence of an electron acceptor, and 5-hydroxy-isoquinoline, which rapidly and very effectively inactivated the enzyme in the presence as well as in the absence of the electron acceptors iodonitrotetrazolium chloride, phenazine methosulfate, or ferricyanide. << Less

  J. Biol. Chem. 269:11254-11260(1994) [PubMed] [EuropePMC]