Cargando…

Production of a Novel Superoxide Dismutase by Escherichia coli and Pichia pastoris and Analysis of the Thermal Stability of the Enzyme

Previously, a new copper-zinc SOD (CuZnSOD) isolated from chestnut rose (Rosa roxburghii) with good stability was described. In this study, the biosynthetic approach was used to create recombinant CuZnSOD. RACE PCR was also used to amplify the full-length CuZnSOD gene from chestnut rose, and the ORF...

Descripción completa

Detalles Bibliográficos
Autores principales: Zhao, Yang, Zhao, Liang, Zhang, Weiwei, Rao, Lei, Wang, Yongtao, Liao, Xiaojun
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8959677/
https://www.ncbi.nlm.nih.gov/pubmed/35356736
http://dx.doi.org/10.3389/fnut.2022.850824
_version_ 1784677214710661120
author Zhao, Yang
Zhao, Liang
Zhang, Weiwei
Rao, Lei
Wang, Yongtao
Liao, Xiaojun
author_facet Zhao, Yang
Zhao, Liang
Zhang, Weiwei
Rao, Lei
Wang, Yongtao
Liao, Xiaojun
author_sort Zhao, Yang
collection PubMed
description Previously, a new copper-zinc SOD (CuZnSOD) isolated from chestnut rose (Rosa roxburghii) with good stability was described. In this study, the biosynthetic approach was used to create recombinant CuZnSOD. RACE PCR was also used to amplify the full-length CuZnSOD gene from chestnut rose, and the ORF segment was expressed in E. coli BL21 and P. pastoris GS115. For characterization, the enzyme was isolated in two steps in E. coli and one step in P. pastoris. The biochemical properties of the two recombinant enzymes were similar, and their optimal reaction pH and temperature were 6.0 and 50°C, respectively. According to molecular dynamics simulation, the CuZnSOD showed high stability from 70 to 90°C, and eight amino acids are important for enzyme thermal stability at high temperatures. This study set the stage for industrial manufacture by filling gaps in the link between conformational changes and the thermal stability of the new CuZnSOD.
format Online
Article
Text
id pubmed-8959677
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher Frontiers Media S.A.
record_format MEDLINE/PubMed
spelling pubmed-89596772022-03-29 Production of a Novel Superoxide Dismutase by Escherichia coli and Pichia pastoris and Analysis of the Thermal Stability of the Enzyme Zhao, Yang Zhao, Liang Zhang, Weiwei Rao, Lei Wang, Yongtao Liao, Xiaojun Front Nutr Nutrition Previously, a new copper-zinc SOD (CuZnSOD) isolated from chestnut rose (Rosa roxburghii) with good stability was described. In this study, the biosynthetic approach was used to create recombinant CuZnSOD. RACE PCR was also used to amplify the full-length CuZnSOD gene from chestnut rose, and the ORF segment was expressed in E. coli BL21 and P. pastoris GS115. For characterization, the enzyme was isolated in two steps in E. coli and one step in P. pastoris. The biochemical properties of the two recombinant enzymes were similar, and their optimal reaction pH and temperature were 6.0 and 50°C, respectively. According to molecular dynamics simulation, the CuZnSOD showed high stability from 70 to 90°C, and eight amino acids are important for enzyme thermal stability at high temperatures. This study set the stage for industrial manufacture by filling gaps in the link between conformational changes and the thermal stability of the new CuZnSOD. Frontiers Media S.A. 2022-03-09 /pmc/articles/PMC8959677/ /pubmed/35356736 http://dx.doi.org/10.3389/fnut.2022.850824 Text en Copyright © 2022 Zhao, Zhao, Zhang, Rao, Wang and Liao. https://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 Nutrition
Zhao, Yang
Zhao, Liang
Zhang, Weiwei
Rao, Lei
Wang, Yongtao
Liao, Xiaojun
Production of a Novel Superoxide Dismutase by Escherichia coli and Pichia pastoris and Analysis of the Thermal Stability of the Enzyme
title Production of a Novel Superoxide Dismutase by Escherichia coli and Pichia pastoris and Analysis of the Thermal Stability of the Enzyme
title_full Production of a Novel Superoxide Dismutase by Escherichia coli and Pichia pastoris and Analysis of the Thermal Stability of the Enzyme
title_fullStr Production of a Novel Superoxide Dismutase by Escherichia coli and Pichia pastoris and Analysis of the Thermal Stability of the Enzyme
title_full_unstemmed Production of a Novel Superoxide Dismutase by Escherichia coli and Pichia pastoris and Analysis of the Thermal Stability of the Enzyme
title_short Production of a Novel Superoxide Dismutase by Escherichia coli and Pichia pastoris and Analysis of the Thermal Stability of the Enzyme
title_sort production of a novel superoxide dismutase by escherichia coli and pichia pastoris and analysis of the thermal stability of the enzyme
topic Nutrition
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8959677/
https://www.ncbi.nlm.nih.gov/pubmed/35356736
http://dx.doi.org/10.3389/fnut.2022.850824
work_keys_str_mv AT zhaoyang productionofanovelsuperoxidedismutasebyescherichiacoliandpichiapastorisandanalysisofthethermalstabilityoftheenzyme
AT zhaoliang productionofanovelsuperoxidedismutasebyescherichiacoliandpichiapastorisandanalysisofthethermalstabilityoftheenzyme
AT zhangweiwei productionofanovelsuperoxidedismutasebyescherichiacoliandpichiapastorisandanalysisofthethermalstabilityoftheenzyme
AT raolei productionofanovelsuperoxidedismutasebyescherichiacoliandpichiapastorisandanalysisofthethermalstabilityoftheenzyme
AT wangyongtao productionofanovelsuperoxidedismutasebyescherichiacoliandpichiapastorisandanalysisofthethermalstabilityoftheenzyme
AT liaoxiaojun productionofanovelsuperoxidedismutasebyescherichiacoliandpichiapastorisandanalysisofthethermalstabilityoftheenzyme