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Identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in Vibrio algivorus

Alginate is a marine non-food-competing polysaccharide that has potential applications in biorefinery. Owing to its large size (molecular weight >300,000 Da), alginate cannot pass through the bacterial cell membrane. Therefore, bacteria that utilize alginate are presumed to have an enzyme that de...

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Autores principales: Doi, Hidetaka, Tokura, Yuriko, Mori, Yukiko, Mori, Kenichi, Asakura, Yoko, Usuda, Yoshihiro, Fukuda, Hiroo, Chinen, Akito
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Berlin Heidelberg 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5266763/
https://www.ncbi.nlm.nih.gov/pubmed/27915375
http://dx.doi.org/10.1007/s00253-016-8021-7
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author Doi, Hidetaka
Tokura, Yuriko
Mori, Yukiko
Mori, Kenichi
Asakura, Yoko
Usuda, Yoshihiro
Fukuda, Hiroo
Chinen, Akito
author_facet Doi, Hidetaka
Tokura, Yuriko
Mori, Yukiko
Mori, Kenichi
Asakura, Yoko
Usuda, Yoshihiro
Fukuda, Hiroo
Chinen, Akito
author_sort Doi, Hidetaka
collection PubMed
description Alginate is a marine non-food-competing polysaccharide that has potential applications in biorefinery. Owing to its large size (molecular weight >300,000 Da), alginate cannot pass through the bacterial cell membrane. Therefore, bacteria that utilize alginate are presumed to have an enzyme that degrades extracellular alginate. Recently, Vibrio algivorus sp. SA2(T) was identified as a novel alginate-decomposing and alginate-utilizing species. However, little is known about the mechanism of alginate degradation and metabolism in this species. To address this issue, we screened the V. algivorus genomic DNA library for genes encoding polysaccharide-decomposing enzymes using a novel double-layer plate screening method and identified alyB as a candidate. Most identified alginate-decomposing enzymes (i.e., alginate lyases) must be concentrated and purified before extracellular alginate depolymerization. AlyB of V. algivorus heterologously expressed in Escherichia coli depolymerized extracellular alginate without requiring concentration or purification. We found seven homologues in the V. algivorus genome (alyB, alyD, oalA, oalB, oalC, dehR, and toaA) that are thought to encode enzymes responsible for alginate transport and metabolism. Introducing these genes into E. coli enabled the cells to assimilate soluble alginate depolymerized by V. algivorus AlyB as the sole carbon source. The alginate was bioconverted into l-lysine (43.3 mg/l) in E. coli strain AJIK01. These findings demonstrate a simple and novel screening method for identifying polysaccharide-degrading enzymes in bacteria and provide a simple alginate biocatalyst and fermentation system with potential applications in industrial biorefinery. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00253-016-8021-7) contains supplementary material, which is available to authorized users.
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spelling pubmed-52667632017-02-09 Identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in Vibrio algivorus Doi, Hidetaka Tokura, Yuriko Mori, Yukiko Mori, Kenichi Asakura, Yoko Usuda, Yoshihiro Fukuda, Hiroo Chinen, Akito Appl Microbiol Biotechnol Applied Genetics and Molecular Biotechnology Alginate is a marine non-food-competing polysaccharide that has potential applications in biorefinery. Owing to its large size (molecular weight >300,000 Da), alginate cannot pass through the bacterial cell membrane. Therefore, bacteria that utilize alginate are presumed to have an enzyme that degrades extracellular alginate. Recently, Vibrio algivorus sp. SA2(T) was identified as a novel alginate-decomposing and alginate-utilizing species. However, little is known about the mechanism of alginate degradation and metabolism in this species. To address this issue, we screened the V. algivorus genomic DNA library for genes encoding polysaccharide-decomposing enzymes using a novel double-layer plate screening method and identified alyB as a candidate. Most identified alginate-decomposing enzymes (i.e., alginate lyases) must be concentrated and purified before extracellular alginate depolymerization. AlyB of V. algivorus heterologously expressed in Escherichia coli depolymerized extracellular alginate without requiring concentration or purification. We found seven homologues in the V. algivorus genome (alyB, alyD, oalA, oalB, oalC, dehR, and toaA) that are thought to encode enzymes responsible for alginate transport and metabolism. Introducing these genes into E. coli enabled the cells to assimilate soluble alginate depolymerized by V. algivorus AlyB as the sole carbon source. The alginate was bioconverted into l-lysine (43.3 mg/l) in E. coli strain AJIK01. These findings demonstrate a simple and novel screening method for identifying polysaccharide-degrading enzymes in bacteria and provide a simple alginate biocatalyst and fermentation system with potential applications in industrial biorefinery. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00253-016-8021-7) contains supplementary material, which is available to authorized users. Springer Berlin Heidelberg 2016-12-03 2017 /pmc/articles/PMC5266763/ /pubmed/27915375 http://dx.doi.org/10.1007/s00253-016-8021-7 Text en © The Author(s) 2016 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.
spellingShingle Applied Genetics and Molecular Biotechnology
Doi, Hidetaka
Tokura, Yuriko
Mori, Yukiko
Mori, Kenichi
Asakura, Yoko
Usuda, Yoshihiro
Fukuda, Hiroo
Chinen, Akito
Identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in Vibrio algivorus
title Identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in Vibrio algivorus
title_full Identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in Vibrio algivorus
title_fullStr Identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in Vibrio algivorus
title_full_unstemmed Identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in Vibrio algivorus
title_short Identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in Vibrio algivorus
title_sort identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in vibrio algivorus
topic Applied Genetics and Molecular Biotechnology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5266763/
https://www.ncbi.nlm.nih.gov/pubmed/27915375
http://dx.doi.org/10.1007/s00253-016-8021-7
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