EFFECTS OF TEMPERATURE AND NOVOBIOCIN ON THE EXPRESSION OF CALF PROCHYMOSIN GENE AND ON PLASMID COPY NUMBER IN RECOMBINANT ESCHERICHIA-COLI

Escherichia coli strain HB101 harboring an expression plasmid bearing calf prochymosin gene under the control of the tac promoter was grown in the presence of IPTG with or without novobiocin at 28 and 40 degree s C, respectively. The differential rates of synthesis of prochymosin inclusions, and, for comparison, of beta-lactamase and beta-galactosid ase, as well as plasmid copy number, were determined during the first hours of steady state growth. At 28 degrees C the induced expression o f prochymosin gene was almost blocked. Addition of novobiocin did not alleviate this effect. In fact, it strenghtened it, and we conclude th at both these additive inhibitory effects are a consequence of the dec rease in negative superhelical tension of plasmid DNA to an insufficie nt level. At 40 degrees C the differential rate of prochymosin synthes is was markedly enhanced. Since the copy number of the expression plas mid increased approximately to the same extent, we conclude that an in crease in gene dose is the cause. The stimulation of cloned heterologo us gene expression at 40 degrees C and inhibition al 28 degrees C may be conveniently used in biotechnological-scale cultivations of some re combinant bacteria.