Inducible high expression of the Escherichia coli infC gene subcloned behind a bacteriophage T7 promoter.
The gene for Escherichia coli translational initiation factor 3 (infC) has been inserted into an overexpression plasmid under the control of the bacteriophage T7 promoter. The infC plasmid was then used to transform a host with a chromosomal T7 RNA polymerase gene controlled by the lacUV5 promoter. Induction of T7 RNA polymerase expression in the host cells resulted in a 200-fold overexpression of infC mRNA and a 100-fold overproduction of initiation factor 3. Rapid batch purification of biologically active IF3 yielded predominantly the long form of IF3, implying that the short form is an artifact of purification by traditional methods.
Muralikrishna
P
P
Department of Chemistry, University of South Florida, Tampa 33620.
Wickstrom
E
E
D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000235:genetics*; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D010448:Peptide Initiation Factors / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D010957:Plasmids; D039665:Prokaryotic Initiation Factor-3; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D013604:T-Phages / Q000235:genetics
D001426:Bacterial Proteins; D004274:DNA, Recombinant; D010448:Peptide Initiation Factors; D039665:Prokaryotic Initiation Factor-3; D012333:RNA, Messenger; D011994:Recombinant Proteins; D007544:Isopropyl Thiogalactoside