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. In order 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 ismore » 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.« less

Authors:

Gu, Wenyu ^[1]; Farhan Ul Haque, Muhammad ^[1]; Baral, Bipin S. ^[2]; Turpin, Erick A. ^[2]; Bandow, Nathan L. ^[2]; Kremmer, Elisabeth ^[3]; Flatley, Andrew ^[3]; Zischka, Hans ^[4]; DiSpirito, Alan A. ^[2]; Semrau, Jeremy D. ^[1]

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+ Show Author Affiliations

1. Univ. of Michigan, Ann Arbor, MI (United States). Dept. of Civil and Environmental Engineering
2. Iowa State Univ., Ames, IA (United States). Roy J. Carver Dept. of Biochemistry, Biophysics and Molecular Biology
3. Helmholtz-Center Munich (HZM) (Germany). German Research Center for Environmental Health (HMGU) and Inst. of Molecular Immunology and Monoclonal Antibody Core Facility
4. Helmholtz-Center Munich (HZM), Neuherberg (Germany). German Research Center for Environmental Health (HMGU) and Inst. of Molecular Toxicology and Pharmacology

Publication Date:

Mon Mar 07 00:00:00 EST 2016

Research Org.:

Univ. of Michigan, Ann Arbor, MI (United States)

Sponsoring Org.:

USDOE Office of Science (SC), Biological and Environmental Research (BER)

OSTI Identifier:

1470736

Grant/Contract Number:  

SC0006630

Resource Type:

Accepted Manuscript

Journal Name:

Applied and Environmental Microbiology

Additional Journal Information:

Journal Volume: 82; Journal Issue: 6; Journal ID: ISSN 0099-2240

Publisher:

American Society for Microbiology

Country of Publication:

United States

Language:

English

Subject:

59 BASIC BIOLOGICAL SCIENCES; 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY