IlvIH Operon of Escherichia coli is Positively Regulated

摘要

The ilvIH operon of Escherichia coli(located near min 2) encodes acetohydroxyacidsynthase III, an isozyme involved in branched-chain amino acid biosynthesis. A strain with lacZ fused to the ilvIH promoter was constructed. Transposon Tn10 was introduced into this strain, and tetracycline-resistant derivatives were screened for those in which ilvIH promoter expression was markedly reduced. In one such derivative, strain CV1008, Beta-galactosidase expression was reduced more than 30-fold. The transposon giving rise to this phenotype inserted near min 20 on the E. coli chromosome. Extract for a wild-type strain contains a protein, the IHB protein, that binds to two sites upstream of the ilvIH promoter (E. Ricca, D.A. Aker, and J.M. Calvo, J. Bacteriol. 171:1658-1664, 1989). Extract from strain CV1008 lacks IHB-binding activity. These results indicate that the IHB protein is a positive regulator of ilvIH operon expression. The gene that encodes the IHB protein, ihb, was cloned by complementing the transposon-induced mutation. Definitive evidence that the cloned DNA encodes the IHB protein was provided by determining the sequence of more than 17 amino acids at the N terminus of the IHB protein and comparing it with the nucleotide sequence. A mutation that prevents repression of the ilvIH operon by leucine in vivo and that alters the DNA-binding characteristics of the IHB protein in vitro was shown to be an allele of the ihb gene. (js)