Molecular cloning and characterisation of the gua regulatory region of Escherichia coli K12

Abstract

The regulatory region of the gua operon of Escherichia coli is contained within a 2.1 kb EcoR1 restriction fragment isolated from a lambda pgua transducing phage. This DNA fragment was inserted into pPV33-II, a promoter-cloning vector, where it activated the gene(s) for tetracycline resistance. The level of tetracycline resistance conferred by the hybrid plasmid was reduced by the addition of guanine and increased by adenine, indicating the presence of the gua promoter. The cloned fragment codes for a polypeptide that complements in vivo the defective enzymes present in certain guaB mutants. This polypeptide was characterised using minicells and immunoprecipitation, and is presumed to correspond to the N-terminal region of IMP dehydrogenase.