Harvest Active Recombinant Rho Kinase from Escherichia coli(Molecular and Cell Biology)

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概要

• 論文の詳細を見る

• Rho kinase (ROCK) inhibitors are effective candidates for treating nerve or myocardial injury, erectile dysfunction, and other cardiovascular diseases. Purified ROCK is a foundation for ROCK inhibitors screening and for its function research in vitro. This article established an easy way to harvest active recombinant ROCK catalytic domain (ROCK-CD) of rat in Escherichia coli (E. coli). The cDNA of ROCK-CD was amplified by RT-PCR, and subcloned to pET28a (+) vector to express the protein in E. coli BL(21) as inclusion bodies. The protein was purified by HiTrap chelating column, and its refolding was achieved by gradient dilution from guanidine hydrochloride solution, and desalinated by ultrafiltration. The result of DNA sequencing and protein sequence analysis indicate there were three amino acid residua of mutation, but the activity was not significantly affected. The activity of the recombinant protein was confirmed by ROCK II activity fluorescence polarization kit. Therefore, this is an easy and rapid procedure to harvest a large quantity of activity recombinant ROCK-CD at low cost.

• 社団法人日本薬学会の論文
• 2006-01-01

著者

• Zhang Luyong

  Jiangsu Center for Drug Screening, China Pharmaceutical University

• Liu Jun

  Key Lab Of New Drugs Design And Discovery Of Liaoning Province School Of Pharmaceutical Engineering

• Duan Weigang

  National Drug Screening Center China Pharmaceutical University

• ZHANG Luyong

  National Drug Screening Center, China Pharmaceutical University

• DUAN WeiGang

  Jiangsu Center for Drug Screening, China Pharmaceutical University

• SUN LiXin

  Jiangsu Center for Drug Screening, China Pharmaceutical University

• LIU Jun

  Jiangsu Center for Drug Screening, China Pharmaceutical University

• YAN Ming

  Jiangsu Center for Drug Screening, China Pharmaceutical University

• WANG Shanzhi

  Jiangsu Center for Drug Screening, China Pharmaceutical University

• CHEN Min

  Jiangsu Center for Drug Screening, China Pharmaceutical University

• WANG Cuifen

  Jiangsu Center for Drug Screening, China Pharmaceutical University

• Yan Ming

  Jiangsu Center For Drug Screening China Pharmaceutical University

• Sun Lixin

  Jiangsu Center For Drug Screening China Pharmaceutical University

• Wang Cuifen

  Jiangsu Center For Drug Screening China Pharmaceutical University

• Wang Shanzhi

  Jiangsu Center For Drug Screening China Pharmaceutical University

• Duan Weigang

  Jiangsu Center For Drug Screening China Pharmaceutical University

• Zhang Luyong

  National Drug Screening Center China Pharmaceutical University

• Zhang Luyong

  Jiangsu Center For Drug Screening China Pharmaceutical University

• Chen Min

  Jiangsu Center For Drug Screening China Pharmaceutical University

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