Regulation of expression of the cloned ada gene in Escherichia coli

Abstract

The ada gene of Escherichia coli K12, the regulatory gene for the adaptive response of bacteria to alkylating agents, was cloned in multicopy plasmids. O6-Methylguanine-DNA methyltransferase and 3-methyladenine-DNA glycosylase II, which are known to be inducible as part of the adaptive response, were produced in ada- cells bearing ada+ plasmids, even without treatment with alkylating agents. When such cells had been treated with methyl methanesulfonate, even higher levels of the enzyme activities were produced. Maxicell experiments revealed that the ada gene codes for a polypeptide with a molecular weight of 38 000. We constructed a hybrid plasmid carrying an ada'-lacZ' fused gene, with the proper control region for ada expression. beta-Galactosidase synthesis from the fused gene was strongly induced only when cells were treated with low doses of methylating agents, but was weakly induced with relatively high doses of ethylating agents. The induction was autogenously regulated by the ada gene product, in a positive manner.