Cloning, expression and purification of pyruvate, orthophosphate dikinase regulatory proteins from <em>Phyllostachys edulis</em>

WANG Chaoli; ZHANG Zhijun; QU Yaping; WANG Lei

doi:10.11833/j.issn.2095-0756.2015.05.014

Volume 32 Issue 5

Sep. 2015

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WANG Chaoli, ZHANG Zhijun, QU Yaping, WANG Lei. Cloning, expression and purification of pyruvate, orthophosphate dikinase regulatory proteins from Phyllostachys edulis[J]. Journal of Zhejiang A&F University, 2015, 32(5): 749-755. doi: 10.11833/j.issn.2095-0756.2015.05.014

Citation:

WANG Chaoli, ZHANG Zhijun, QU Yaping, WANG Lei. Cloning, expression and purification of pyruvate, orthophosphate dikinase regulatory proteins from Phyllostachys edulis[J]. Journal of Zhejiang A&F University, 2015, 32(5): 749-755. doi: 10.11833/j.issn.2095-0756.2015.05.014

Cloning, expression and purification of pyruvate, orthophosphate dikinase regulatory proteins from Phyllostachys edulis

doi: 10.11833/j.issn.2095-0756.2015.05.014

1.
The Nurturing Station for the State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Lin’an 311300, Zhejiang, China

Funds:

国家自然科学基金资助项目（31270715）;浙江省自然科学基金资助项目（LY14C160009）

Received Date: 2014-11-28
Rev Recd Date: 2015-01-09
Publish Date: 2015-10-20

Abstract

Pyruvate orthophosphate dikinase regulatory protein （PPDK-RP） is a key protein in C4 photosynthesis and can modified enzyme activity of PPDK. For Phyllostachys edulis, an important economic bamboo species, cloning PPDK-RP gene and studying its functions had vital theoretical value and application for bamboo photosynthesis research. Firstly PeRP1 was successfully cloned by reverse transcription polymerase chain reaction （RT-PCR）, and afterward a multiple sequence alignment and phylogenetic analysis was conducted. Then for further study the crystal structure of the PeRP protein, the recombinant expression pe-SUMO vectors of PeRP were constructed and the protein was expressed in E. coli and purified by nickle beads column and size column. Results showed that the PeRP gene contained a 1.275 kb open reading frame （ORF） coding a 425 amino acid polypeptide, which contained the typical domain of UDF299 and was predicted as a number of kinase-PPPase superfamily. The multiple sequence alignment and phylogenetic analysis of PeRP revealed that Phyllostachys edulis was a typical C3 monocotyledon. Expressed soluble PeRP1 protein had three forms--polymer, dimmers, and monomers in water soultion. These provided a foundation for the structure and function of RP.［Ch, 7 fig. 15 ref.］
- botany,
- Phyllostachys edulis,
- pyruvate, orthophosphate dikinase regulatory proteins,
- gene cloning,
- prokaryotic expression,
- protein purification

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

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Cloning, expression and purification of pyruvate, orthophosphate dikinase regulatory proteins from Phyllostachys edulis

1. The Nurturing Station for the State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Lin’an 311300, Zhejiang, China

Funds:

国家自然科学基金资助项目（31270715）;浙江省自然科学基金资助项目（LY14C160009）

Received Date: 2014-11-28

Rev Recd Date: 2015-01-09

Publish Date: 2015-10-20

Key words:

botany /
Phyllostachys edulis /
pyruvate, orthophosphate dikinase regulatory proteins /
gene cloning /
prokaryotic expression /
protein purification

Abstract: Pyruvate orthophosphate dikinase regulatory protein （PPDK-RP） is a key protein in C4 photosynthesis and can modified enzyme activity of PPDK. For Phyllostachys edulis, an important economic bamboo species, cloning PPDK-RP gene and studying its functions had vital theoretical value and application for bamboo photosynthesis research. Firstly PeRP1 was successfully cloned by reverse transcription polymerase chain reaction （RT-PCR）, and afterward a multiple sequence alignment and phylogenetic analysis was conducted. Then for further study the crystal structure of the PeRP protein, the recombinant expression pe-SUMO vectors of PeRP were constructed and the protein was expressed in E. coli and purified by nickle beads column and size column. Results showed that the PeRP gene contained a 1.275 kb open reading frame （ORF） coding a 425 amino acid polypeptide, which contained the typical domain of UDF299 and was predicted as a number of kinase-PPPase superfamily. The multiple sequence alignment and phylogenetic analysis of PeRP revealed that Phyllostachys edulis was a typical C3 monocotyledon. Expressed soluble PeRP1 protein had three forms--polymer, dimmers, and monomers in water soultion. These provided a foundation for the structure and function of RP.［Ch, 7 fig. 15 ref.］

Cloning, expression and purification of pyruvate, orthophosphate dikinase regulatory proteins from Phyllostachys edulis

doi: 10.11833/j.issn.2095-0756.2015.05.014

Abstract

References

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通讯作者: 陈斌, bchen63@163.com

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Cloning, expression and purification of pyruvate, orthophosphate dikinase regulatory proteins from Phyllostachys edulis

doi: 10.11833/j.issn.2095-0756.2015.05.014

1. The Nurturing Station for the State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Lin’an 311300, Zhejiang, China

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Cloning, expression and purification of pyruvate, orthophosphate dikinase regulatory proteins from Phyllostachys edulis

doi: 10.11833/j.issn.2095-0756.2015.05.014

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通讯作者: 陈斌, bchen63@163.com

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Cloning, expression and purification of pyruvate, orthophosphate dikinase regulatory proteins from Phyllostachys edulis

doi: 10.11833/j.issn.2095-0756.2015.05.014

1. The Nurturing Station for the State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Lin’an 311300, Zhejiang, China

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