Cloning and promoter analysis of theEscherichia coliadenylate cyclase gene

Abstract

The gene for adenylate cyclase of E. coli has been cloned in the plasmid pBR322. The Cya- strain transformed with the isolated plasmids produces significant amounts of adenylate cyclase and cAMP. Some of the Cya+ plasmids were shown to direct the synthesis of a 85,000 dalton polypeptide in a cell-free system. The direction of transcription and the location of the cya promoter including the transcriptional start site were determined by an S1 digestion method. DNA sequence around the promoter region indicates that a putative coding region for adenylate cyclase begins at +233. The 233 bp leader region could encode a potential small polypeptide containing 30 amino acids. Two probable CRP binding sites were found in the leader region, suggesting a negative control at the transcriptional level by CRP-cAMP.