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Removal of N-terminal tail changes the thermostability of the low-temperature-active exo-inulinase InuAGN25

Exo-inulinases are members of the glycoside hydrolase family 32 and function by hydrolyzing inulin into fructose with yields up to 90–95%. The N-terminal tail contributes to enzyme thermotolerance, which plays an important role in enzyme applications. However, the role of N-terminal amino acid resid...

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Autores principales: He, Limei, Zhang, Rui, Shen, Jidong, Miao, Ying, Tang, Xianghua, Wu, Qian, Zhou, Junpei, Huang, Zunxi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Taylor & Francis 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8291819/
https://www.ncbi.nlm.nih.gov/pubmed/32865156
http://dx.doi.org/10.1080/21655979.2020.1809921
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author He, Limei
Zhang, Rui
Shen, Jidong
Miao, Ying
Tang, Xianghua
Wu, Qian
Zhou, Junpei
Huang, Zunxi
author_facet He, Limei
Zhang, Rui
Shen, Jidong
Miao, Ying
Tang, Xianghua
Wu, Qian
Zhou, Junpei
Huang, Zunxi
author_sort He, Limei
collection PubMed
description Exo-inulinases are members of the glycoside hydrolase family 32 and function by hydrolyzing inulin into fructose with yields up to 90–95%. The N-terminal tail contributes to enzyme thermotolerance, which plays an important role in enzyme applications. However, the role of N-terminal amino acid residues in the thermal performance and structural properties of exo-inulinases remains to be elucidated. In this study, three and six residues of the N-terminus starting from Gln23 of the exo-inulinase InuAGN25 were deleted and expressed in Escherichia coli. After digestion with human rhinovirus 3 C protease to remove the N-terminal amino acid fusion sequence that may affect the thermolability of enzymes, wild-type RfsMInuAGN25 and its mutants RfsMutNGln23Δ3 and RfsMutNGln23Δ6 were produced. Compared with RfsMInuAGN25, thermostability of RfsMutNGln23Δ3 was enhanced while that of RfsMutNGln23Δ6 was slightly reduced. Compared with the N-terminal structures of RfsMInuAGN25 and RfsMutNGln23Δ6, RfsMutNGln23Δ3 had a higher content of (1) the helix structure, (2) salt bridges (three of which were organized in a network), (3) cation–π interactions (one of which anchored the N-terminal tail). These structural properties may account for the improved thermostability of RfsMutNGln23Δ3. The study provides a better understanding of the N-terminus–function relationships that are useful for rational design of thermostability of exo-inulinases.
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spelling pubmed-82918192021-08-30 Removal of N-terminal tail changes the thermostability of the low-temperature-active exo-inulinase InuAGN25 He, Limei Zhang, Rui Shen, Jidong Miao, Ying Tang, Xianghua Wu, Qian Zhou, Junpei Huang, Zunxi Bioengineered Research Paper Exo-inulinases are members of the glycoside hydrolase family 32 and function by hydrolyzing inulin into fructose with yields up to 90–95%. The N-terminal tail contributes to enzyme thermotolerance, which plays an important role in enzyme applications. However, the role of N-terminal amino acid residues in the thermal performance and structural properties of exo-inulinases remains to be elucidated. In this study, three and six residues of the N-terminus starting from Gln23 of the exo-inulinase InuAGN25 were deleted and expressed in Escherichia coli. After digestion with human rhinovirus 3 C protease to remove the N-terminal amino acid fusion sequence that may affect the thermolability of enzymes, wild-type RfsMInuAGN25 and its mutants RfsMutNGln23Δ3 and RfsMutNGln23Δ6 were produced. Compared with RfsMInuAGN25, thermostability of RfsMutNGln23Δ3 was enhanced while that of RfsMutNGln23Δ6 was slightly reduced. Compared with the N-terminal structures of RfsMInuAGN25 and RfsMutNGln23Δ6, RfsMutNGln23Δ3 had a higher content of (1) the helix structure, (2) salt bridges (three of which were organized in a network), (3) cation–π interactions (one of which anchored the N-terminal tail). These structural properties may account for the improved thermostability of RfsMutNGln23Δ3. The study provides a better understanding of the N-terminus–function relationships that are useful for rational design of thermostability of exo-inulinases. Taylor & Francis 2020-08-30 /pmc/articles/PMC8291819/ /pubmed/32865156 http://dx.doi.org/10.1080/21655979.2020.1809921 Text en © 2020 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group. https://creativecommons.org/licenses/by/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) ), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research Paper
He, Limei
Zhang, Rui
Shen, Jidong
Miao, Ying
Tang, Xianghua
Wu, Qian
Zhou, Junpei
Huang, Zunxi
Removal of N-terminal tail changes the thermostability of the low-temperature-active exo-inulinase InuAGN25
title Removal of N-terminal tail changes the thermostability of the low-temperature-active exo-inulinase InuAGN25
title_full Removal of N-terminal tail changes the thermostability of the low-temperature-active exo-inulinase InuAGN25
title_fullStr Removal of N-terminal tail changes the thermostability of the low-temperature-active exo-inulinase InuAGN25
title_full_unstemmed Removal of N-terminal tail changes the thermostability of the low-temperature-active exo-inulinase InuAGN25
title_short Removal of N-terminal tail changes the thermostability of the low-temperature-active exo-inulinase InuAGN25
title_sort removal of n-terminal tail changes the thermostability of the low-temperature-active exo-inulinase inuagn25
topic Research Paper
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8291819/
https://www.ncbi.nlm.nih.gov/pubmed/32865156
http://dx.doi.org/10.1080/21655979.2020.1809921
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