title: A TonB-Dependent Transporter Is Responsible for Methanobactin Uptake by Methylosinus trichosporium OB3b.
abstract: Methanobactin, a small modified polypeptide synthesized by methanotrophs for copper uptake, has been found to be chromosomally encoded. The gene encoding the polypeptide precursor of methanobactin, mbnA, is part of a gene cluster that also includes several genes encoding proteins of unknown function (but speculated to be involved in methanobactin formation) as well as mbnT, which encodes a TonB-dependent transporter hypothesized to be responsible for methanobactin uptake. To determine if mbnT is truly responsible for methanobactin uptake, a knockout was constructed in Methylosinus trichosporium OB3b using marker exchange mutagenesis. The resulting M. trichosporium mbnT::Gm(r) mutant was found to be able to produce methanobactin but was unable to internalize it. Further, if this mutant was grown in the presence of copper and exogenous methanobactin, copper uptake was significantly reduced. Expression of mmoX and pmoA, encoding polypeptides of the soluble methane monooxygenase (sMMO) and particulate methane monooxygenase (pMMO), respectively, also changed significantly when methanobactin was added, which indicates that the mutant was unable to collect copper under these conditions. Copper uptake and gene expression, however, were not affected in wild-type M. trichosporium OB3b, indicating that the TonB-dependent transporter encoded by mbnT is responsible for methanobactin uptake and that methanobactin is a key mechanism used by methanotrophs for copper uptake. When the mbnT::Gm(r) mutant was grown under a range of copper concentrations in the absence of methanobactin, however, the phenotype of the mutant was indistinguishable from that of wild-type M. trichosporium OB3b, indicating that this methanotroph has multiple mechanisms for copper uptake.
authors.affiliation: Department of Civil and Environmental Engineering, University of Michigan, Ann Arbor, Michigan, USA.
authors.forename: Wenyu
authors.initials: W
authors.lastname: Gu
authors.affiliation: Department of Civil and Environmental Engineering, University of Michigan, Ann Arbor, Michigan, USA.
authors.forename: Muhammad
authors.initials: M
authors.lastname: Farhan Ul Haque
authors.affiliation: Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, Iowa, USA.
authors.forename: Bipin S
authors.initials: BS
authors.lastname: Baral
authors.affiliation: Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, Iowa, USA.
authors.forename: Erick A
authors.initials: EA
authors.lastname: Turpin
authors.affiliation: Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, Iowa, USA.
authors.forename: Nathan L
authors.initials: NL
authors.lastname: Bandow
authors.affiliation: Institute of Molecular Immunology, Core Facility Monoclonal Antibodies, Helmholtz Center Munich, German Research Center for Environmental Health, Munich, Germany.
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Kremmer
authors.affiliation: Institute of Molecular Immunology, Core Facility Monoclonal Antibodies, Helmholtz Center Munich, German Research Center for Environmental Health, Munich, Germany.
authors.forename: Andrew
authors.initials: A
authors.lastname: Flatley
authors.affiliation: Institute of Molecular Toxicology and Pharmacology, Helmholtz Center Munich, German Research Center for Environmental Health, Neuherberg, Germany.
authors.forename: Hans
authors.initials: H
authors.lastname: Zischka
authors.affiliation: Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, Iowa, USA.
authors.forename: Alan A
authors.initials: AA
authors.lastname: DiSpirito
authors.affiliation: Department of Civil and Environmental Engineering, University of Michigan, Ann Arbor, Michigan, USA jsemrau@umich.edu.
authors.forename: Jeremy D
authors.initials: JD
authors.lastname: Semrau
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D003300:Copper / Q000378:metabolism; D055786:Gene Knockout Techniques; D007093:Imidazoles / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D020602:Methylosinus trichosporium / Q000235:genetics / Q000378:metabolism*; D009842:Oligopeptides / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D007093:Imidazoles; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D009842:Oligopeptides; C491147:methanobactin; C057616:tonB protein, Bacteria; D003300:Copper