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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...

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Autores principales: 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
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2020
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.
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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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