THERMO-INDUCIBLE EXPRESSION OF A RECOMBINANT FUSION PROTEIN BY ESCHERICHIA-COLI LAC REPRESSOR MUTANTS

The lac I gene of Escherichia coli encodes the lactose repressor. We i solated temperature-sensitive mutants of the lac I gene by in vitro mu tagenesis with hydroxylamine. The new mutation sites were determined, and replacement of a single amino acid had respectively occurred at am ino acid positions 241 (Ala -->Thr), 265 (Gly --> Asp) and 300 (Ser -- > Asn). These mutation sites were located in the core region of the in c repressor protein. Temperature-dependent expression of beta-galactos idase was observed in the strains having these mutant lac I genes. By using these temperature-sensitive lac I genes, we developed a thermo-i nducible expression system for a foreign gene under the control of the lac promoter. A recombinant fusion protein, consisting of a derivativ e of E. coli beta-galactosidase and the human calcitonin precursor pep tide, was efficiently produced by using this system.