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Discovery and Biochemical Characterization of a Methanol Dehydrogenase From Lysinibacillus xylanilyticus
Bioconversion of C1 chemicals such as methane and methanol into higher carbon-chain chemicals has been widely studied. Methanol oxidation catalyzed by methanol dehydrogenase (Mdh) is one of the key steps in methanol utilization in bacterial methylotrophy. In bacteria, few NAD(+)-dependent Mdhs have...
Autores principales: | , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Frontiers Media S.A.
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7033420/ https://www.ncbi.nlm.nih.gov/pubmed/32117944 http://dx.doi.org/10.3389/fbioe.2020.00067 |
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author | Lee, Jin-Young Park, Sung-Hyun Oh, So-Hyung Lee, Jin-Ju Kwon, Kil Koang Kim, Su-Jin Choi, Minjeong Rha, Eugene Lee, Hyewon Lee, Dae-Hee Sung, Bong Hyun Yeom, Soo-Jin Lee, Seung-Goo |
author_facet | Lee, Jin-Young Park, Sung-Hyun Oh, So-Hyung Lee, Jin-Ju Kwon, Kil Koang Kim, Su-Jin Choi, Minjeong Rha, Eugene Lee, Hyewon Lee, Dae-Hee Sung, Bong Hyun Yeom, Soo-Jin Lee, Seung-Goo |
author_sort | Lee, Jin-Young |
collection | PubMed |
description | Bioconversion of C1 chemicals such as methane and methanol into higher carbon-chain chemicals has been widely studied. Methanol oxidation catalyzed by methanol dehydrogenase (Mdh) is one of the key steps in methanol utilization in bacterial methylotrophy. In bacteria, few NAD(+)-dependent Mdhs have been reported that convert methanol to formaldehyde. In this study, an uncharacterized Mdh gene from Lysinibacillus xylanilyticus (Lxmdh) was cloned and expressed in Escherichia coli. The maximum alcohol oxidation activity of the recombinant enzyme was observed at pH 9.5 and 55°C in the presence of 10 mM Mg(2+). To improve oxidation activity, rational approach-based, site-directed mutagenesis of 16 residues in the putative active site and NAD(+)-binding region was performed. The mutations S101V, T141S, and A164F improved the enzyme’s specific activity toward methanol compared to that of the wild-type enzyme. These mutants show a slightly higher turnover rate than that of wild-type, although their K(M) values were increased compared to that of wild-type. Consequently, according the kinetic results, S101, T141, and A164 positions may related to the catalytic activity in the active site for methanol dehydrogenation. It should be further studied other mutant variants with high activity for methanol. In conclusion, we characterized a new Lxmdh and its variants that may be potentially useful for the development of synthetic methylotrophy in the future. |
format | Online Article Text |
id | pubmed-7033420 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-70334202020-02-28 Discovery and Biochemical Characterization of a Methanol Dehydrogenase From Lysinibacillus xylanilyticus Lee, Jin-Young Park, Sung-Hyun Oh, So-Hyung Lee, Jin-Ju Kwon, Kil Koang Kim, Su-Jin Choi, Minjeong Rha, Eugene Lee, Hyewon Lee, Dae-Hee Sung, Bong Hyun Yeom, Soo-Jin Lee, Seung-Goo Front Bioeng Biotechnol Bioengineering and Biotechnology Bioconversion of C1 chemicals such as methane and methanol into higher carbon-chain chemicals has been widely studied. Methanol oxidation catalyzed by methanol dehydrogenase (Mdh) is one of the key steps in methanol utilization in bacterial methylotrophy. In bacteria, few NAD(+)-dependent Mdhs have been reported that convert methanol to formaldehyde. In this study, an uncharacterized Mdh gene from Lysinibacillus xylanilyticus (Lxmdh) was cloned and expressed in Escherichia coli. The maximum alcohol oxidation activity of the recombinant enzyme was observed at pH 9.5 and 55°C in the presence of 10 mM Mg(2+). To improve oxidation activity, rational approach-based, site-directed mutagenesis of 16 residues in the putative active site and NAD(+)-binding region was performed. The mutations S101V, T141S, and A164F improved the enzyme’s specific activity toward methanol compared to that of the wild-type enzyme. These mutants show a slightly higher turnover rate than that of wild-type, although their K(M) values were increased compared to that of wild-type. Consequently, according the kinetic results, S101, T141, and A164 positions may related to the catalytic activity in the active site for methanol dehydrogenation. It should be further studied other mutant variants with high activity for methanol. In conclusion, we characterized a new Lxmdh and its variants that may be potentially useful for the development of synthetic methylotrophy in the future. Frontiers Media S.A. 2020-02-14 /pmc/articles/PMC7033420/ /pubmed/32117944 http://dx.doi.org/10.3389/fbioe.2020.00067 Text en Copyright © 2020 Lee, Park, Oh, Lee, Kwon, Kim, Choi, Rha, Lee, Lee, Sung, Yeom and Lee. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Bioengineering and Biotechnology Lee, Jin-Young Park, Sung-Hyun Oh, So-Hyung Lee, Jin-Ju Kwon, Kil Koang Kim, Su-Jin Choi, Minjeong Rha, Eugene Lee, Hyewon Lee, Dae-Hee Sung, Bong Hyun Yeom, Soo-Jin Lee, Seung-Goo Discovery and Biochemical Characterization of a Methanol Dehydrogenase From Lysinibacillus xylanilyticus |
title | Discovery and Biochemical Characterization of a Methanol Dehydrogenase From Lysinibacillus xylanilyticus |
title_full | Discovery and Biochemical Characterization of a Methanol Dehydrogenase From Lysinibacillus xylanilyticus |
title_fullStr | Discovery and Biochemical Characterization of a Methanol Dehydrogenase From Lysinibacillus xylanilyticus |
title_full_unstemmed | Discovery and Biochemical Characterization of a Methanol Dehydrogenase From Lysinibacillus xylanilyticus |
title_short | Discovery and Biochemical Characterization of a Methanol Dehydrogenase From Lysinibacillus xylanilyticus |
title_sort | discovery and biochemical characterization of a methanol dehydrogenase from lysinibacillus xylanilyticus |
topic | Bioengineering and Biotechnology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7033420/ https://www.ncbi.nlm.nih.gov/pubmed/32117944 http://dx.doi.org/10.3389/fbioe.2020.00067 |
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