Identification of a two-component regulatory pathway essential for Mn(II) oxidation in pseudomonas putida GB-1

Kati Geszvain, Bradley Tebo

Research output: Contribution to journalArticle

35 Citations (Scopus)

Abstract

Bacterial manganese (II) oxidation has a profound impact on the biogeochemical cycling of Mn and the availability of the trace metals adsorbed to the surfaces of solid Mn(III, IV) oxides. The Mn(II) oxidase enzyme was tentatively identified in Pseudomonas putida GB-I via transposon mutagenesis: the mutant strain GB-1-007, which fails to oxidize Mn(II), harbors a transposon insertion in the gene cumA. cumA encodes a putative multicopper oxidase (MCO), a class of enzymes implicated in Mn(II) oxidation in other bacterial species. However, we show here that an in-frame deletion of cumA did not affect Mn(II) oxidation. Through complementation analysis of the oxidation defect in GB-1-007 with a cosmid library and subsequent sequencing of candidate genes we show the causative mutation to be a frameshift within the mnxSl gene that encodes a putative sensor histidine kinase. The frameshift mutation results in a truncated protein lacking the kinase domain. Multicopy expression ofmnxSl restored Mn(II) oxidation to GB-1-007 and in-frame deletion of mnxSl resulted in a loss of oxidation in the wild-type strain. These results clearly demonstrated that the oxidation defect of GB-1-007 is due to disruption of mnxSl, not cumA Tn5, and that CumA is not the Mn(II) oxidase. mnxSl is located upstream of a second sensor histidine kinase gene, mnxS2, and a response regulator gene, mnxR. In-frame deletions of each of these genes also led to the loss of Mn(II) oxidation. Therefore, we conclude that the MnxS1/MnxS2/MnxR two-component regulatory pathway is essential for Mn(II) oxidation in P. putida GB-1.

Original languageEnglish (US)
Pages (from-to)1224-1231
Number of pages8
JournalApplied and Environmental Microbiology
Volume76
Issue number4
DOIs
StatePublished - Feb 2010

Fingerprint

Pseudomonas putida
Oxidoreductases
oxidation
Genes
Cosmids
Frameshift Mutation
gene
Insertional Mutagenesis
Gene Deletion
Enzymes
Regulator Genes
Manganese
Mutagenesis
Oxides
Protein Kinases
histidine kinase
Metals
genes
transposons
Mutation

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Food Science
  • Biotechnology
  • Ecology

Cite this

Identification of a two-component regulatory pathway essential for Mn(II) oxidation in pseudomonas putida GB-1. / Geszvain, Kati; Tebo, Bradley.

In: Applied and Environmental Microbiology, Vol. 76, No. 4, 02.2010, p. 1224-1231.

Research output: Contribution to journalArticle

@article{75cf69382989431d8c7502b2893fcdb0,
title = "Identification of a two-component regulatory pathway essential for Mn(II) oxidation in pseudomonas putida GB-1",
abstract = "Bacterial manganese (II) oxidation has a profound impact on the biogeochemical cycling of Mn and the availability of the trace metals adsorbed to the surfaces of solid Mn(III, IV) oxides. The Mn(II) oxidase enzyme was tentatively identified in Pseudomonas putida GB-I via transposon mutagenesis: the mutant strain GB-1-007, which fails to oxidize Mn(II), harbors a transposon insertion in the gene cumA. cumA encodes a putative multicopper oxidase (MCO), a class of enzymes implicated in Mn(II) oxidation in other bacterial species. However, we show here that an in-frame deletion of cumA did not affect Mn(II) oxidation. Through complementation analysis of the oxidation defect in GB-1-007 with a cosmid library and subsequent sequencing of candidate genes we show the causative mutation to be a frameshift within the mnxSl gene that encodes a putative sensor histidine kinase. The frameshift mutation results in a truncated protein lacking the kinase domain. Multicopy expression ofmnxSl restored Mn(II) oxidation to GB-1-007 and in-frame deletion of mnxSl resulted in a loss of oxidation in the wild-type strain. These results clearly demonstrated that the oxidation defect of GB-1-007 is due to disruption of mnxSl, not cumA Tn5, and that CumA is not the Mn(II) oxidase. mnxSl is located upstream of a second sensor histidine kinase gene, mnxS2, and a response regulator gene, mnxR. In-frame deletions of each of these genes also led to the loss of Mn(II) oxidation. Therefore, we conclude that the MnxS1/MnxS2/MnxR two-component regulatory pathway is essential for Mn(II) oxidation in P. putida GB-1.",
author = "Kati Geszvain and Bradley Tebo",
year = "2010",
month = "2",
doi = "10.1128/AEM.02473-09",
language = "English (US)",
volume = "76",
pages = "1224--1231",
journal = "Applied and Environmental Microbiology",
issn = "0099-2240",
publisher = "American Society for Microbiology",
number = "4",

}

TY - JOUR

T1 - Identification of a two-component regulatory pathway essential for Mn(II) oxidation in pseudomonas putida GB-1

AU - Geszvain, Kati

AU - Tebo, Bradley

PY - 2010/2

Y1 - 2010/2

N2 - Bacterial manganese (II) oxidation has a profound impact on the biogeochemical cycling of Mn and the availability of the trace metals adsorbed to the surfaces of solid Mn(III, IV) oxides. The Mn(II) oxidase enzyme was tentatively identified in Pseudomonas putida GB-I via transposon mutagenesis: the mutant strain GB-1-007, which fails to oxidize Mn(II), harbors a transposon insertion in the gene cumA. cumA encodes a putative multicopper oxidase (MCO), a class of enzymes implicated in Mn(II) oxidation in other bacterial species. However, we show here that an in-frame deletion of cumA did not affect Mn(II) oxidation. Through complementation analysis of the oxidation defect in GB-1-007 with a cosmid library and subsequent sequencing of candidate genes we show the causative mutation to be a frameshift within the mnxSl gene that encodes a putative sensor histidine kinase. The frameshift mutation results in a truncated protein lacking the kinase domain. Multicopy expression ofmnxSl restored Mn(II) oxidation to GB-1-007 and in-frame deletion of mnxSl resulted in a loss of oxidation in the wild-type strain. These results clearly demonstrated that the oxidation defect of GB-1-007 is due to disruption of mnxSl, not cumA Tn5, and that CumA is not the Mn(II) oxidase. mnxSl is located upstream of a second sensor histidine kinase gene, mnxS2, and a response regulator gene, mnxR. In-frame deletions of each of these genes also led to the loss of Mn(II) oxidation. Therefore, we conclude that the MnxS1/MnxS2/MnxR two-component regulatory pathway is essential for Mn(II) oxidation in P. putida GB-1.

AB - Bacterial manganese (II) oxidation has a profound impact on the biogeochemical cycling of Mn and the availability of the trace metals adsorbed to the surfaces of solid Mn(III, IV) oxides. The Mn(II) oxidase enzyme was tentatively identified in Pseudomonas putida GB-I via transposon mutagenesis: the mutant strain GB-1-007, which fails to oxidize Mn(II), harbors a transposon insertion in the gene cumA. cumA encodes a putative multicopper oxidase (MCO), a class of enzymes implicated in Mn(II) oxidation in other bacterial species. However, we show here that an in-frame deletion of cumA did not affect Mn(II) oxidation. Through complementation analysis of the oxidation defect in GB-1-007 with a cosmid library and subsequent sequencing of candidate genes we show the causative mutation to be a frameshift within the mnxSl gene that encodes a putative sensor histidine kinase. The frameshift mutation results in a truncated protein lacking the kinase domain. Multicopy expression ofmnxSl restored Mn(II) oxidation to GB-1-007 and in-frame deletion of mnxSl resulted in a loss of oxidation in the wild-type strain. These results clearly demonstrated that the oxidation defect of GB-1-007 is due to disruption of mnxSl, not cumA Tn5, and that CumA is not the Mn(II) oxidase. mnxSl is located upstream of a second sensor histidine kinase gene, mnxS2, and a response regulator gene, mnxR. In-frame deletions of each of these genes also led to the loss of Mn(II) oxidation. Therefore, we conclude that the MnxS1/MnxS2/MnxR two-component regulatory pathway is essential for Mn(II) oxidation in P. putida GB-1.

UR - http://www.scopus.com/inward/record.url?scp=76649094256&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=76649094256&partnerID=8YFLogxK

U2 - 10.1128/AEM.02473-09

DO - 10.1128/AEM.02473-09

M3 - Article

VL - 76

SP - 1224

EP - 1231

JO - Applied and Environmental Microbiology

JF - Applied and Environmental Microbiology

SN - 0099-2240

IS - 4

ER -