title: Distal end of 105-125 loop--a putative reductase binding domain of phthalate dioxygenase.
abstract: The phthalate dioxygenase system consists of the dioxygenase, PDO, which contains a Rieske [2Fe-2S] center and a Fe(II)-mononuclear center, and the reductase, PDR. Involvement of the distal end of the 105-125 loop of PDO in its interaction with PDR was tested by substituting charged residues in the loop with alanines and by replacing the conserved tryptophan-94. Compared to wild-type PDO, all variants had lower catalytic activity and the Rieske centers were reduced more slowly by reduced PDR. The rates of oxidation of the Rieske centers by oxygen, which represent electron transfer between the Rieske and mononuclear centers, were essentially unaffected. These results suggest that positively charged residues of the distal end of the 105-125 loop are collectively involved in PDR binding with the PDO. Contrary to expectations, Trp94 variants were not directly involved in electron transfer between PDR and PDO. The tryptophan appears to have mainly a structural role, apparently preserving the hydrophilic environment of the Rieske center.
authors.affiliation: Dept. of Biological Chemistry, University of Michigan, 1150 W. Medical Center Dr., MSRBIII, Ann Arbor, MI 48109-0606, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Tarasev
authors.forename: Sailaja
authors.initials: S
authors.lastname: Pullela
authors.forename: David P
authors.initials: DP
authors.lastname: Ballou
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D001665:Binding Sites; D016956:Burkholderia cepacia / Q000201:enzymology / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D004579:Electron Transport; D014450:Electron Transport Complex III / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D010088:Oxidoreductases / Q000737:chemistry / Q000378:metabolism; D010105:Oxygenases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial; D009097:Multienzyme Complexes; D011994:Recombinant Proteins; C037989:Rieske iron-sulfur protein; D010088:Oxidoreductases; C049135:phthalate oxygenase reductase; D010105:Oxygenases; C044732:phthalate dioxygenase; D014450:Electron Transport Complex III