Enhancing recombinant protein yields in Escherichia coli using the T7 system under the control of heat inducible lambda PL promoter.
A recombinant plasmid containing the complete lacZ gene downstream of the T7 promoter was used to transform Escherichia coli containing another plasmid which had the T7 RNA polymerase gene under the control of heat inducible lambda PL promoter. This recombinant E. coli containing the two plasmids was studied in order to enhance beta-galactosidase expression. The heat shock time which effectively regulates the T7 RNA polymerase was optimized and best expression of beta-galactosidase was obtained with 2 min heat shock. Substrate feeding increased the duration of log phase and allowed induction at a higher cell density without affecting the specific activity. A high cell density (7 g l-1) and high specific activity (approximately 20,000 U) were achieved which effectively increased the product concentration 18-fold.
Gupta
J C
JC
Centre for Biotechnology, Jawaharlal Nehru University, New Delhi, India.
Jaisani
M
M
Pandey
G
G
Mukherjee
K J
KJ
D017123:Bacteriophage T7 / Q000235:genetics*; D003470:Culture Media; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D018869:Heat-Shock Response; D006358:Hot Temperature; D007763:Lac Operon; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis*; D001616:beta-Galactosidase / Q000378:metabolism
D003470:Culture Media; D011994:Recombinant Proteins; D012321:DNA-Directed RNA Polymerases; D001616:beta-Galactosidase