Mutagenesis and regulation of the cysJ promoter of Escherichia coli K-12.
The cysJ promoter of Escherichia coli K-12, which is positively controlled by the CysB regulatory protein, was localized through the formation of a fusion of cysJ, the gene encoding NADPH-cytochrome c reductase with lacZ. The position of the transcription start point was determined and the orientation of transcription was shown to be counterclockwise on the E. coli K-12 map. Oligodeoxyribonucleotide-directed mutagenesis of the inferred -10 and -35 regions indicated that both sites could be altered to produce promoter 'down' mutations. When the -10 region was made to agree with the -10 consensus sequence, there was increased function under conditions of repression (that is, in the presence of cysteine). Upstream deletions, as well as mutations in a region proposed to be involved in binding of the CysB regulatory protein, identified sequences important for promoter activity from -90 to -78 and from -71 to -66. By comparison of the sequences of four cys promoters, a possible CysB-binding site was found which included the region shown to be required for the positive regulation of the cysJ promoter.
Loudon
J A
JA
Department of Biochemistry, University of Sydney, NSW, Australia.
Loughlin
R E
RE
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050862:Oxidoreductases Acting on Sulfur Group Donors / Q000235:genetics*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012689:Sequence Homology, Nucleic Acid; D050864:Sulfite Reductase (NADPH)
D001426:Bacterial Proteins; C062886:CysB protein, Bacteria; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D050862:Oxidoreductases Acting on Sulfur Group Donors; D050864:Sulfite Reductase (NADPH); C494314:sulfite reductase (NADPH), E coli