An efficient cell-free protein synthesis system using periplasmic phosphatase-removed S30 extract

• Published in: Journal of microbiological methods Vol. 43; no. 2; pp. 91 - 96
• Main Authors: Kang, Sang Hyeon, Oh, Taek Jin, Kim, Ryang Guk, Kang, Taek Jin, Hwang, Seung Ha, Lee, Eun Yeol, Choi, Cha Yong
• Format: Journal Article
• Language: English
• Published: Shannon Elsevier B.V 15.12.2000; Elsevier Science
• Subjects: Biological and medical sciences; Biotechnology; Cell-Free System; Cell-free translation; Escherichia coli - metabolism; Fundamental and applied biological sciences. Psychology; Methods. Procedures. Technologies; Periplasm - enzymology; Phosphatase; Phosphoric Monoester Hydrolases - metabolism; Protein Biosynthesis; Protein engineering; Spheroplast; Spheroplasts - metabolism; Subcellular Fractions - metabolism; Phosphatase; Cell-free translation; Spheroplast; In vitro translation; Enzyme; Protein synthesis; Escherichia coli; Phosphoric monoester hydrolases; Bacteria; Hydrolases; Esterases; Genetic translation; Enterobacteriaceae
• ISSN: 0167-7012; 1872-8359
• DOI: 10.1016/S0167-7012(00)00206-2

An efficient cell-free translation system was developed by removal of phosphatase localized in the periplasmic space, which hampers the translation reaction by hydrolyzing ATP. S30 extract was prepared from the spheroplast of Escherichia coli, and as much as 40% of ATP-hydrolysis activity of phosphatases could be easily removed by the spheroplast formation. The reaction period of translation using phosphatase-removed S30 extract could be prolonged and protein synthesis was enhanced by more than 30%.