Expression, Purification, and Characterization of a Sucrose Nonfermenting 1-Related Protein Kinases 2 of Arabidopsis thaliana in E. coli-Based Cell-Free System

The plant-specific sucrose nonfermenting 1-related protein kinase 2 (SnRK2) family is considered an important regulator of plant responses to abiotic stresses such as drought, cold, salinity, and nutrition deficiency. However, little information is available on how SnRK2s regulate sulfur deprivation...

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Main Authors: Xu Zhang, Shuangxi Zhang, Jun Wang, Jing Zi, Jianhui Wang, Shaolin Chen, Yi Wan
Format: Article
Language:English
Published: Hindawi Limited 2016-01-01
Series:BioMed Research International
Online Access:http://dx.doi.org/10.1155/2016/9469356
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spelling doaj-54a4e9f3bea34f4b8dcd98be090c72622020-11-24T23:10:43ZengHindawi LimitedBioMed Research International2314-61332314-61412016-01-01201610.1155/2016/94693569469356Expression, Purification, and Characterization of a Sucrose Nonfermenting 1-Related Protein Kinases 2 of Arabidopsis thaliana in E. coli-Based Cell-Free SystemXu Zhang0Shuangxi Zhang1Jun Wang2Jing Zi3Jianhui Wang4Shaolin Chen5Yi Wan6Microbiology Institute of Shaanxi, Shaanxi Academy of Sciences, 76 Xiying Road, Xi’an, Shaanxi 710043, ChinaBiomass Energy Center for Arid and Semiarid Lands, College of Life Sciences, Northwest A&F University, 3 Taicheng Road, Yangling, Shaanxi 712100, ChinaMicrobiology Institute of Shaanxi, Shaanxi Academy of Sciences, 76 Xiying Road, Xi’an, Shaanxi 710043, ChinaMicrobiology Institute of Shaanxi, Shaanxi Academy of Sciences, 76 Xiying Road, Xi’an, Shaanxi 710043, ChinaMicrobiology Institute of Shaanxi, Shaanxi Academy of Sciences, 76 Xiying Road, Xi’an, Shaanxi 710043, ChinaBiomass Energy Center for Arid and Semiarid Lands, College of Life Sciences, Northwest A&F University, 3 Taicheng Road, Yangling, Shaanxi 712100, ChinaMicrobiology Institute of Shaanxi, Shaanxi Academy of Sciences, 76 Xiying Road, Xi’an, Shaanxi 710043, ChinaThe plant-specific sucrose nonfermenting 1-related protein kinase 2 (SnRK2) family is considered an important regulator of plant responses to abiotic stresses such as drought, cold, salinity, and nutrition deficiency. However, little information is available on how SnRK2s regulate sulfur deprivation responses in Arabidopsis. Large-scale production of SnRK2 kinases in vitro can help to elucidate the biochemical properties and physiological functions of this protein family. However, heterogenous expression of SnRK2s usually leads to inactive proteins. In this study, we expressed a recombinant Arabidopsis SnRK2.1 in a modified E. coli cell-free system, which combined two kinds of extracts allowing for a convenient and affordable protein preparation. The recombinant SnRK2.1 was produced in large-scale and the autophosphorylation activity of purified SnRK2.1 was characterized, allowing for further biochemical and substrate binding analysis in sulfur signaling. The application of this improved E. coli cell-free system provides us a promising and convenient platform to enhance expression of the target proteins economically.http://dx.doi.org/10.1155/2016/9469356
collection DOAJ
language English
format Article
sources DOAJ
author Xu Zhang
Shuangxi Zhang
Jun Wang
Jing Zi
Jianhui Wang
Shaolin Chen
Yi Wan
spellingShingle Xu Zhang
Shuangxi Zhang
Jun Wang
Jing Zi
Jianhui Wang
Shaolin Chen
Yi Wan
Expression, Purification, and Characterization of a Sucrose Nonfermenting 1-Related Protein Kinases 2 of Arabidopsis thaliana in E. coli-Based Cell-Free System
BioMed Research International
author_facet Xu Zhang
Shuangxi Zhang
Jun Wang
Jing Zi
Jianhui Wang
Shaolin Chen
Yi Wan
author_sort Xu Zhang
title Expression, Purification, and Characterization of a Sucrose Nonfermenting 1-Related Protein Kinases 2 of Arabidopsis thaliana in E. coli-Based Cell-Free System
title_short Expression, Purification, and Characterization of a Sucrose Nonfermenting 1-Related Protein Kinases 2 of Arabidopsis thaliana in E. coli-Based Cell-Free System
title_full Expression, Purification, and Characterization of a Sucrose Nonfermenting 1-Related Protein Kinases 2 of Arabidopsis thaliana in E. coli-Based Cell-Free System
title_fullStr Expression, Purification, and Characterization of a Sucrose Nonfermenting 1-Related Protein Kinases 2 of Arabidopsis thaliana in E. coli-Based Cell-Free System
title_full_unstemmed Expression, Purification, and Characterization of a Sucrose Nonfermenting 1-Related Protein Kinases 2 of Arabidopsis thaliana in E. coli-Based Cell-Free System
title_sort expression, purification, and characterization of a sucrose nonfermenting 1-related protein kinases 2 of arabidopsis thaliana in e. coli-based cell-free system
publisher Hindawi Limited
series BioMed Research International
issn 2314-6133
2314-6141
publishDate 2016-01-01
description The plant-specific sucrose nonfermenting 1-related protein kinase 2 (SnRK2) family is considered an important regulator of plant responses to abiotic stresses such as drought, cold, salinity, and nutrition deficiency. However, little information is available on how SnRK2s regulate sulfur deprivation responses in Arabidopsis. Large-scale production of SnRK2 kinases in vitro can help to elucidate the biochemical properties and physiological functions of this protein family. However, heterogenous expression of SnRK2s usually leads to inactive proteins. In this study, we expressed a recombinant Arabidopsis SnRK2.1 in a modified E. coli cell-free system, which combined two kinds of extracts allowing for a convenient and affordable protein preparation. The recombinant SnRK2.1 was produced in large-scale and the autophosphorylation activity of purified SnRK2.1 was characterized, allowing for further biochemical and substrate binding analysis in sulfur signaling. The application of this improved E. coli cell-free system provides us a promising and convenient platform to enhance expression of the target proteins economically.
url http://dx.doi.org/10.1155/2016/9469356
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