title: Elimination of manganese(II,III) oxidation in Pseudomonas putida GB-1 by a double knockout of two putative multicopper oxidase genes.
abstract: Bacterial manganese(II) oxidation impacts the redox cycling of Mn, other elements, and compounds in the environment; therefore, it is important to understand the mechanisms of and enzymes responsible for Mn(II) oxidation. In several Mn(II)-oxidizing organisms, the identified Mn(II) oxidase belongs to either the multicopper oxidase (MCO) or the heme peroxidase family of proteins. However, the identity of the oxidase in Pseudomonas putida GB-1 has long remained unknown. To identify the P. putida GB-1 oxidase, we searched its genome and found several homologues of known or suspected Mn(II) oxidase-encoding genes (mnxG, mofA, moxA, and mopA). To narrow this list, we assumed that the Mn(II) oxidase gene would be conserved among Mn(II)-oxidizing pseudomonads but not in nonoxidizers and performed a genome comparison to 11 Pseudomonas species. We further assumed that the oxidase gene would be regulated by MnxR, a transcription factor required for Mn(II) oxidation. Two loci met all these criteria: PputGB1_2447, which encodes an MCO homologous to MnxG, and PputGB1_2665, which encodes an MCO with very low homology to MofA. In-frame deletions of each locus resulted in strains that retained some ability to oxidize Mn(II) or Mn(III); loss of oxidation was attained only upon deletion of both genes. These results suggest that PputGB1_2447 and PputGB1_2665 encode two MCOs that are independently capable of oxidizing both Mn(II) and Mn(III). The purpose of this redundancy is unclear; however, differences in oxidation phenotype for the single mutants suggest specialization in function for the two enzymes.
authors.affiliation: Division of Environmental and Biomolecular Systems, Institute of Environmental Health, Oregon Health & Science University, Beaverton, Oregon, USA. geszvaik@ebs.ogi.edu
authors.forename: Kati
authors.initials: K
authors.lastname: Geszvain
authors.forename: James K
authors.initials: JK
authors.lastname: McCarthy
authors.forename: Bradley M
authors.initials: BM
authors.lastname: Tebo
mesh_terms: D055786:Gene Knockout Techniques; D008345:Manganese / Q000378:metabolism*; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000235:genetics*; D016958:Pseudomonas putida / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*
chemical_list: D008345:Manganese; D010088:Oxidoreductases