Effects of mutations altering SOS regulation on a nalidixic acid-inducible system for the production of heterologous proteins inEscherichia coli

The major leftward early promoter of phage λpL, has frequently been used to drive expression of heterologous genes inEscherichia coli.pL is typically maintained fully repressed by the lambda cl protein. When induction of heterologous protein synthesis is desired, one of several potential mechanisms of destroying cl function is employed and the expression of the foreign gene commences. One method of derepressingpL involves exposing cells to nalidixic acid, which results in the “activation” of RecA protein and the subsequent RecA-mediated proteolytic cleavage of cl. Activated RecA also mediates the cleavage of theE. coli LexA protein, resulting in induction of the SOS regulon (at least 15E. coli genes, includingrec A). We have examined the effect of two chromosomal mutations on the productivity of nalidixic acid inductions. One of the tested mutations (recA o) increased the intracellular concentration of RecA prior to induction; the other (lexAind−) resulted in a mutated lexA protein insensitive to RecA-mediated cleavage. These mutations were introduced into a strain carrying acl+ defective lysogen. Synthesis of two heterologous proteins, human α1-antitrypsin and a fusion protein partially derived from thePlasmodium falciparum circumsporozooite surface antigen, was examined in the wild-type and mutant strains. The maximum α-1 antitrypsin concentration achieved was improved by 50% when therecA o strain was used rather than the wild type; however; only smaller changes (20% or less) in the maximum concentration of the malaria fusion protein wer observed. Use of thelexAind− strain resulted in a decrease in the maximum concentration attained for both heterologous products.