Repression of Escherichia coli purB is by a transcriptional roadblock mechanism.
Escherichia coli purB is regulated by a repressor-operator interaction. The purB operator is 242 bp downstream from the transcription start site and overlaps condons 62 to 67 in the protein-coding sequence (B. He, J. M. Smith, and H. Zalkin, J. Bacteriol. 174:130-136, 1992). The mechanism by which the repressor-operator interaction functions to repress transcription was investigated by a combination of promoter replacement experiments and RNA analyses. By using a trp promoter replacement that deleted 5' flanking DNA to position -986, purB expression was increased sevenfold, yet normal two- to threefold regulation was maintained. This indicates that repressor-operator control is independent of the purB promoter and other 5' flanking sequences. Transcriptional regulation was likewise independent of coupled translation. An approximately 260-nucleotide truncated in vivo purB mRNA was identified which was dependent upon repressor-operator interaction. Thus, binding of purine repressor to the purB operator inhibits transcription elongation by a roadblock mechanism. The roadblock was not influenced by a sevenfold increase in promoter strength or by an operator mutation resulting in a 2.5-fold increase in repressor-operator affinity.
He
B
B
Department of Biochemistry, Purdue University, West Lafayette, Indiana 47907.
Zalkin
H
H
D001483:Base Sequence; D003001:Cloning, Molecular; D003062:Codon; D004926:Escherichia coli / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D012097:Repressor Proteins / Q000378:metabolism; D015183:Restriction Mapping; D014158:Transcription, Genetic; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D003062:Codon; D009838:Oligodeoxyribonucleotides; D012097:Repressor Proteins; D001616:beta-Galactosidase