Heterologous protein production in Escherichia coli using the propionate-inducible pPro system by conventional and auto-induction methods.
We examined expression of two plant genes encoding coclaurine N-methyltransferase (CMT) and norcoclaurine synthase (NCS) in Escherichia coli from the Salmonella entericaprpBCDE promoter (P(prpB)) and compared it to that from the strongest IPTG-inducible promoter, P(T7). In contrast to our previous study showing slightly higher production of green fluorescent protein (GFP) from the pPro system compared to that from the T7 system, production of two plant proteins CMT and NCS from P(prpB) was 2- to 4-fold higher than that from P(T7). Unlike P(T7), expression from P(prpB) did not reduce cell growth even when highly induced, indicating that this propionate-inducible system is more efficient for overproduction of proteins that result in growth inhibition. In an auto-induction experiment, which does not require monitoring the culture or adding inducer during cell growth, the pPro system exhibited much higher protein production than the T7 system. These results strongly indicate that the pPro system is well-suited for overproduction of recombinant proteins.
Lee
Sung Kuk
SK
Department of Chemical Engineering, University of California, Berkeley, CA 94720, USA. sungkuk@berkeley.edu
Keasling
Jay D
JD
D019731:Carbon-Nitrogen Ligases / Q000096:biosynthesis* / Q000187:drug effects / Q000235:genetics; D049109:Cell Proliferation / Q000187:drug effects; D003470:Culture Media / Q000494:pharmacology; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D008780:Methyltransferases / Q000096:biosynthesis* / Q000187:drug effects / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011422:Propionates / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics; D013997:Time Factors; D015533:Transcriptional Activation / Q000187:drug effects
D003470:Culture Media; D011422:Propionates; D011994:Recombinant Proteins; D007544:Isopropyl Thiogalactoside; D008780:Methyltransferases; C430217:S-adenosyl-L-methionine coclaurine N-methyltransferase; D019731:Carbon-Nitrogen Ligases; C450500:norcoclaurine synthase