An Aminotransferase is Responsible for the Deamination of the N-terminal Leucine and Required for Formation of Oxazolone Ring A in Methanobactin of Methylosinus trichosporium OB3b
Abstract
Gene expression in methanotrophs has been shown to be affected by the availability of a variety of metals, most notably copper regulating expression of alternative forms of methane monooxygenase. A copper-binding compound or chalkophore called methanobactin plays a key role in copper uptake in methanotrophs. Methanobactin is a ribosomally synthesized and post-translationally modified peptide (RiPP) with two heterocyclic rings with an associated thioamide for each ring, formed from X-Cys dipeptide sequences that bind copper. The gene encoding for the precursor polypeptide of methanobactin,mbnA, is part of a gene cluster, but the role of other genes in methanobactin biosynthesis is unclear. To begin to elucidate the function of these genes, we constructed an unmarked deletion ofmbnABCMNinMethylosinus trichosporiumOB3b, and then homologously expressedmbnABCMoff a broad host range cloning vector to determine the function ofmbnN, annotated as encoding for an aminotransferase. Methanobactin produced by this strain was found to be substantially different from wildtype methanobactin in that the C-terminal methionine was missing, and only one of the two oxazolone rings was formed. Rather, in the place of the N-terminal 3-methylbutanoyl-oxazolone-thioamide group, a leucine and thioamide-containing glycine (Gly-Ψ) were found, indicating that MbnN is used for deamination of the N-terminal leucine of methanobactin, and thatmore »
- Authors:
-
- Univ. of Michigan, Ann Arbor, MI (United States)
- Iowa State Univ., Ames, IA (United States)
- Publication Date:
- Research Org.:
- Univ. of Michigan, Ann Arbor, MI (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER)
- OSTI Identifier:
- 1425406
- Alternate Identifier(s):
- OSTI ID: 1465357
- Grant/Contract Number:
- SC0006630
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Applied and Environmental Microbiology
- Additional Journal Information:
- Journal Volume: 83; Journal Issue: 1; Journal ID: ISSN 0099-2240
- Publisher:
- American Society for Microbiology
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES
Citation Formats
Gu, Wenyu, Baral, Bipin S., DiSpirito, Alan A., and Semrau, Jeremy D. An Aminotransferase is Responsible for the Deamination of the N-terminal Leucine and Required for Formation of Oxazolone Ring A in Methanobactin of Methylosinus trichosporium OB3b. United States: N. p., 2016.
Web. doi:10.1128/AEM.02619-16.
Gu, Wenyu, Baral, Bipin S., DiSpirito, Alan A., & Semrau, Jeremy D. An Aminotransferase is Responsible for the Deamination of the N-terminal Leucine and Required for Formation of Oxazolone Ring A in Methanobactin of Methylosinus trichosporium OB3b. United States. https://doi.org/10.1128/AEM.02619-16
Gu, Wenyu, Baral, Bipin S., DiSpirito, Alan A., and Semrau, Jeremy D. Fri .
"An Aminotransferase is Responsible for the Deamination of the N-terminal Leucine and Required for Formation of Oxazolone Ring A in Methanobactin of Methylosinus trichosporium OB3b". United States. https://doi.org/10.1128/AEM.02619-16. https://www.osti.gov/servlets/purl/1425406.
@article{osti_1425406,
title = {An Aminotransferase is Responsible for the Deamination of the N-terminal Leucine and Required for Formation of Oxazolone Ring A in Methanobactin of Methylosinus trichosporium OB3b},
author = {Gu, Wenyu and Baral, Bipin S. and DiSpirito, Alan A. and Semrau, Jeremy D.},
abstractNote = {Gene expression in methanotrophs has been shown to be affected by the availability of a variety of metals, most notably copper regulating expression of alternative forms of methane monooxygenase. A copper-binding compound or chalkophore called methanobactin plays a key role in copper uptake in methanotrophs. Methanobactin is a ribosomally synthesized and post-translationally modified peptide (RiPP) with two heterocyclic rings with an associated thioamide for each ring, formed from X-Cys dipeptide sequences that bind copper. The gene encoding for the precursor polypeptide of methanobactin,mbnA, is part of a gene cluster, but the role of other genes in methanobactin biosynthesis is unclear. To begin to elucidate the function of these genes, we constructed an unmarked deletion ofmbnABCMNinMethylosinus trichosporiumOB3b, and then homologously expressedmbnABCMoff a broad host range cloning vector to determine the function ofmbnN, annotated as encoding for an aminotransferase. Methanobactin produced by this strain was found to be substantially different from wildtype methanobactin in that the C-terminal methionine was missing, and only one of the two oxazolone rings was formed. Rather, in the place of the N-terminal 3-methylbutanoyl-oxazolone-thioamide group, a leucine and thioamide-containing glycine (Gly-Ψ) were found, indicating that MbnN is used for deamination of the N-terminal leucine of methanobactin, and that this post-translational modification is critical for closure of the N-terminal oxazolone ring inM. trichosporiumOB3b. These studies provide new insights into methanobactin biosynthesis, and also provide a platform for understanding the function of other genes in the methanobactin gene cluster. IMPORTANCEMethanotrophs, microbes that play a critical role in the carbon cycle, are influenced by copper, with gene expression and enzyme activity changing as copper levels change. Methanotrophs produce a copper-binding compound, or chalkophore called methanobactin for copper uptake, and methanobactin plays a key role in controlling methanotrophic activity. Methanobactin has also been shown to be effective in the treatment of Wilson Disease, an autosomal recessive disorder where the human body cannot correctly assimilate copper. It is important that the biosynthetic pathway of methanobactin be characterized to understand how methanotrophs respond to their environment as well as to optimize the use of methanobactin for the treatment of copper-related diseases such as Wilson Disease. Here we show thatmbnN, encoding an aminotransferase, is involved in the deamination of the N-terminal leucine and necessary for the formation of one but not both heterocyclic rings in methanobactin that are responsible for copper binding.},
doi = {10.1128/AEM.02619-16},
journal = {Applied and Environmental Microbiology},
number = 1,
volume = 83,
place = {United States},
year = {Fri Oct 21 00:00:00 EDT 2016},
month = {Fri Oct 21 00:00:00 EDT 2016}
}
Web of Science
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