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Isolation of highly thermostable β-xylosidases from a hot spring soil microbial community using a metagenomic approach

The DNA extracted from a high-temperature environment in which micro-organisms are living will be a good source for the isolation of thermostable enzymes. Using a metagenomic approach, we aimed to isolate thermostable β-xylosidases that will be exploited for biofuel production from lignocellulosic b...

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Autores principales: Sato, Masaru, Suda, Migiwa, Okuma, Jiro, Kato, Tomohiko, Hirose, Yoshitsugu, Nishimura, Asuka, Kondo, Yasuhiko, Shibata, Daisuke
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5726482/
https://www.ncbi.nlm.nih.gov/pubmed/29106502
http://dx.doi.org/10.1093/dnares/dsx032
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author Sato, Masaru
Suda, Migiwa
Okuma, Jiro
Kato, Tomohiko
Hirose, Yoshitsugu
Nishimura, Asuka
Kondo, Yasuhiko
Shibata, Daisuke
author_facet Sato, Masaru
Suda, Migiwa
Okuma, Jiro
Kato, Tomohiko
Hirose, Yoshitsugu
Nishimura, Asuka
Kondo, Yasuhiko
Shibata, Daisuke
author_sort Sato, Masaru
collection PubMed
description The DNA extracted from a high-temperature environment in which micro-organisms are living will be a good source for the isolation of thermostable enzymes. Using a metagenomic approach, we aimed to isolate thermostable β-xylosidases that will be exploited for biofuel production from lignocellulosic biomass. DNA samples obtained from the soil near a spout of a hot spring (70°C, pH7.2) were subjected to sequencing, which generated a total of 84.2 Gbp with 967,925 contigs of >500 bp in length. Similarity search for β-xylosidase in the contigs revealed the presence of 168 candidate sequences, each of which may have arisen from more than one gene. Individual genes were amplified by PCR using sequence-specific primers. The resultant DNA fragments were cloned and introduced into Escherichia coli BL21 Star(DE3). Consequently, 269 proteins were successfully expressed in the E. coli cells and then examined for β-xylosidase activity. A total of 82 proteins exhibited β-xylosidase activity at 50°C, six of which retained the activity even at 90°C. Out of the six, three proteins were originated from a single candidate sequence, AR19M-311. An amino acid sequence comparison suggested the amino acid residues that appeared to be crucial for thermal stability of the enzymes.
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spelling pubmed-57264822017-12-18 Isolation of highly thermostable β-xylosidases from a hot spring soil microbial community using a metagenomic approach Sato, Masaru Suda, Migiwa Okuma, Jiro Kato, Tomohiko Hirose, Yoshitsugu Nishimura, Asuka Kondo, Yasuhiko Shibata, Daisuke DNA Res Full Papers The DNA extracted from a high-temperature environment in which micro-organisms are living will be a good source for the isolation of thermostable enzymes. Using a metagenomic approach, we aimed to isolate thermostable β-xylosidases that will be exploited for biofuel production from lignocellulosic biomass. DNA samples obtained from the soil near a spout of a hot spring (70°C, pH7.2) were subjected to sequencing, which generated a total of 84.2 Gbp with 967,925 contigs of >500 bp in length. Similarity search for β-xylosidase in the contigs revealed the presence of 168 candidate sequences, each of which may have arisen from more than one gene. Individual genes were amplified by PCR using sequence-specific primers. The resultant DNA fragments were cloned and introduced into Escherichia coli BL21 Star(DE3). Consequently, 269 proteins were successfully expressed in the E. coli cells and then examined for β-xylosidase activity. A total of 82 proteins exhibited β-xylosidase activity at 50°C, six of which retained the activity even at 90°C. Out of the six, three proteins were originated from a single candidate sequence, AR19M-311. An amino acid sequence comparison suggested the amino acid residues that appeared to be crucial for thermal stability of the enzymes. Oxford University Press 2017-12 2017-08-07 /pmc/articles/PMC5726482/ /pubmed/29106502 http://dx.doi.org/10.1093/dnares/dsx032 Text en © The Author 2017. Published by Oxford University Press on behalf of Kazusa DNA Research Institute. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Full Papers
Sato, Masaru
Suda, Migiwa
Okuma, Jiro
Kato, Tomohiko
Hirose, Yoshitsugu
Nishimura, Asuka
Kondo, Yasuhiko
Shibata, Daisuke
Isolation of highly thermostable β-xylosidases from a hot spring soil microbial community using a metagenomic approach
title Isolation of highly thermostable β-xylosidases from a hot spring soil microbial community using a metagenomic approach
title_full Isolation of highly thermostable β-xylosidases from a hot spring soil microbial community using a metagenomic approach
title_fullStr Isolation of highly thermostable β-xylosidases from a hot spring soil microbial community using a metagenomic approach
title_full_unstemmed Isolation of highly thermostable β-xylosidases from a hot spring soil microbial community using a metagenomic approach
title_short Isolation of highly thermostable β-xylosidases from a hot spring soil microbial community using a metagenomic approach
title_sort isolation of highly thermostable β-xylosidases from a hot spring soil microbial community using a metagenomic approach
topic Full Papers
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5726482/
https://www.ncbi.nlm.nih.gov/pubmed/29106502
http://dx.doi.org/10.1093/dnares/dsx032
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