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Site-directed Mutagenesis of a β-Glycoside Hydrolase from Lentinula edodes

The β-glycoside hydrolases (LXYL-P1−1 and LXYL-P1−2) from Lentinula edodes (strain M95.33) can specifically hydrolyze 7-β-xylosyl-10-deacetyltaxol (XDT) to form 10-deacetyltaxol for the semi-synthesis of Taxol. Our previous study showed that both the I368T mutation in LXYL-P1−1 and the T368E mutatio...

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Autores principales: Chen, Jing-Jing, Liang, Xiao, Chen, Tian-Jiao, Yang, Jin-Ling, Zhu, Ping
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6337304/
https://www.ncbi.nlm.nih.gov/pubmed/30586935
http://dx.doi.org/10.3390/molecules24010059
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author Chen, Jing-Jing
Liang, Xiao
Chen, Tian-Jiao
Yang, Jin-Ling
Zhu, Ping
author_facet Chen, Jing-Jing
Liang, Xiao
Chen, Tian-Jiao
Yang, Jin-Ling
Zhu, Ping
author_sort Chen, Jing-Jing
collection PubMed
description The β-glycoside hydrolases (LXYL-P1−1 and LXYL-P1−2) from Lentinula edodes (strain M95.33) can specifically hydrolyze 7-β-xylosyl-10-deacetyltaxol (XDT) to form 10-deacetyltaxol for the semi-synthesis of Taxol. Our previous study showed that both the I368T mutation in LXYL-P1−1 and the T368E mutation in LXYL-P1−2 could increase the enzyme activity, which prompted us to investigate the effect of the I368E mutation on LXYL-P1−1 activity. In this study, the β-xylosidase and β-glucosidase activities of LXYL-P1−1(I368E) were 1.5 and 2.2 times higher than those of LXYL-P1−1. Most importantly, combination of I368E and V91S exerted the cumulative effects on the improvement of the enzyme activities and catalytic efficiency. The β-xylosidase and β-glucosidase activities of the double mutant LXYL-P1−1(V91S/I368E) were 3.2 and 1.7-fold higher than those of LXYL-P1−1(I368E). Similarly, the catalytic efficiency of LXYL-P1−1(V91S/I368E) on 7-β-xylosyl-10-deacetyltaxol was 1.8-fold higher than that of LXYL-P1−1(I368E) due to the dramatic increase in the substrate affinity. Molecular docking results suggest that the V91S and I368E mutation might positively promote the interaction between enzyme and substrate through altering the loop conformation near XDT and increasing the hydrogen bonds among Ser(91), Trp(301), and XDT. This study lays the foundation for exploring the relationship between the structure and function of the β-glycoside hydrolases.
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spelling pubmed-63373042019-01-25 Site-directed Mutagenesis of a β-Glycoside Hydrolase from Lentinula edodes Chen, Jing-Jing Liang, Xiao Chen, Tian-Jiao Yang, Jin-Ling Zhu, Ping Molecules Article The β-glycoside hydrolases (LXYL-P1−1 and LXYL-P1−2) from Lentinula edodes (strain M95.33) can specifically hydrolyze 7-β-xylosyl-10-deacetyltaxol (XDT) to form 10-deacetyltaxol for the semi-synthesis of Taxol. Our previous study showed that both the I368T mutation in LXYL-P1−1 and the T368E mutation in LXYL-P1−2 could increase the enzyme activity, which prompted us to investigate the effect of the I368E mutation on LXYL-P1−1 activity. In this study, the β-xylosidase and β-glucosidase activities of LXYL-P1−1(I368E) were 1.5 and 2.2 times higher than those of LXYL-P1−1. Most importantly, combination of I368E and V91S exerted the cumulative effects on the improvement of the enzyme activities and catalytic efficiency. The β-xylosidase and β-glucosidase activities of the double mutant LXYL-P1−1(V91S/I368E) were 3.2 and 1.7-fold higher than those of LXYL-P1−1(I368E). Similarly, the catalytic efficiency of LXYL-P1−1(V91S/I368E) on 7-β-xylosyl-10-deacetyltaxol was 1.8-fold higher than that of LXYL-P1−1(I368E) due to the dramatic increase in the substrate affinity. Molecular docking results suggest that the V91S and I368E mutation might positively promote the interaction between enzyme and substrate through altering the loop conformation near XDT and increasing the hydrogen bonds among Ser(91), Trp(301), and XDT. This study lays the foundation for exploring the relationship between the structure and function of the β-glycoside hydrolases. MDPI 2018-12-24 /pmc/articles/PMC6337304/ /pubmed/30586935 http://dx.doi.org/10.3390/molecules24010059 Text en © 2018 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Chen, Jing-Jing
Liang, Xiao
Chen, Tian-Jiao
Yang, Jin-Ling
Zhu, Ping
Site-directed Mutagenesis of a β-Glycoside Hydrolase from Lentinula edodes
title Site-directed Mutagenesis of a β-Glycoside Hydrolase from Lentinula edodes
title_full Site-directed Mutagenesis of a β-Glycoside Hydrolase from Lentinula edodes
title_fullStr Site-directed Mutagenesis of a β-Glycoside Hydrolase from Lentinula edodes
title_full_unstemmed Site-directed Mutagenesis of a β-Glycoside Hydrolase from Lentinula edodes
title_short Site-directed Mutagenesis of a β-Glycoside Hydrolase from Lentinula edodes
title_sort site-directed mutagenesis of a β-glycoside hydrolase from lentinula edodes
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6337304/
https://www.ncbi.nlm.nih.gov/pubmed/30586935
http://dx.doi.org/10.3390/molecules24010059
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