Enhanced Cell-Free Protein Synthesis Using a S30 Extract from Escherichia coli Grown Rapidly at 42 °C in an Amino Acid Enriched Medium

2008 ◽  
Vol 21 (2) ◽  
pp. 608-613 ◽  
Author(s):  
Tsuneo Yamane ◽  
Yusuke Ikeda ◽  
Takehiro Nagasaka ◽  
Hideo Nakano
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Free Protein ◽  
S30 Extract ◽  
Cell Free Protein Synthesis

scholarly journals Optimizing Cell-Free Protein Synthesis for Increased Yield and Activity of Colicins

2019 ◽  
Vol 2 (2) ◽  
pp. 28 ◽  
Author(s):  
Xing Jin ◽  
Weston Kightlinger ◽  
Seok Hoon Hong
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Cell Killing ◽  
Great Promise ◽  
Immunity Protein ◽  
Free Protein ◽  
Killing Activity ◽  
Colicin E1 ◽  
Colicin E3 ◽  
Cell Free Protein Synthesis

Colicins are antimicrobial proteins produced by Escherichia coli that hold great promise as viable complements or alternatives to antibiotics. Cell-free protein synthesis (CFPS) is a useful production platform for toxic proteins because it eliminates the need to maintain cell viability, a common problem in cell-based production. Previously, we demonstrated that colicins produced by CFPS based on crude Escherichia coli lysates are effective in eradicating antibiotic-tolerant bacteria known as persisters. However, we also found that some colicins have poor solubility or low cell-killing activity. In this study, we improved the solubility of colicin M from 16% to nearly 100% by producing it in chaperone-enriched E. coli extracts, resulting in enhanced cell-killing activity. We also improved the cytotoxicity of colicin E3 by adding or co-expressing the E3 immunity protein during the CFPS reaction, suggesting that the E3 immunity protein enhances colicin E3 activity in addition to protecting the host strain. Finally, we confirmed our previous finding that active colicins can be rapidly synthesized by observing colicin E1 production over time in CFPS. Within three hours of CFPS incubation, colicin E1 reached its maximum production yield and maintained high cytotoxicity during longer incubations up to 20 h. Taken together, our findings indicate that colicin production can be easily optimized for improved solubility and activity using the CFPS platform.

scholarly journals A Highly Efficient Cell-Free Protein Synthesis System from Escherichia coli

1996 ◽  
Vol 239 (3) ◽  
pp. 881-886 ◽  
Author(s):  
Dong-Myung Kim ◽  
Takanori Kigawa ◽  
Cha-Yong Choi ◽  
Shigeyuki Yokoyama
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Synthesis System ◽  
Free Protein ◽  
Highly Efficient ◽  
Protein Synthesis System ◽  
Cell Free Protein Synthesis
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