In vivo evidence that the nusA and infB genes of E. coli are part of the same multi-gene operon which encodes at least four proteins

Abstract

Previous work has shown that the Escherichia coli nusA gene codes for a protein which regulates transcription termination. The 16.0-kb EcoRI DNA fragment that includes the nusA gene, codes for at least eight bacterial proteins of mol. wts. 48 000 (argG), 21 000 (p21), 64 000 (nusA), 120 000 (IF2 alpha)-(infB), 91 000 (IF2 beta)(infB), 15 000 (p15), 10 000 (rpsO) and 85 000 (pnp). We have constructed several deletion and fusion derivatives from this cloned DNA and examined in vivo the structure and expression of these genes. First, the promoter functional in vivo for the nusA gene was mapped at approximately 800 bp upstream of the nusA structural gene. Second, the synthesis of five proteins, p21, NusA, IF2 alpha, IF2 beta (and p15) proteins, was affected by the deletion of the nusA promoter. Third, these same five proteins were hyperproduced after fusion of the DNA fragment to the lambda pL promoter. In addition, subcloning experiments revealed that the p15 gene is expressed by the read-through transcription from the infB gene. These results lead us to conclude that the genes coding for the p21, NusA, InfB (IF2 alpha and IF2 beta), and p15 proteins form a single-transcriptional unit ('nusA-infB operon') in vivo and that rpsO and pnp genes do not belong to the same operon. The in vivo attenuation site of this operon is described.