The Streptomyces galP1 promoter has a novel RNA polymerase recognition sequence and is transcribed by a new form of RNA polymerase in vitro.
We report the identification of DNA sequences that determine the activity of the Streptomyces galP1 promoter and a new form of RNA polymerase holoenzyme that recognizes these sequences in vitro. Base substitutions were introduced throughout the galP1 promoter region, and bases at positions -34, -36, and -11 with respect to the transcription start site were shown to be required for promoter function. These bases correspond in their positions to regions known to be important for RNA polymerase binding in several classes of eubacterial promoters, but the sequences themselves are not similar to those previously described. The -35 region of the galP1 promoter consists of six G residues, and base changes in this G hexamer had a dramatic effect on promoter activity. By using galP1-containing DNA template, a new RNA polymerase activity was purified from Streptomyces. Holoenzyme reconstitution experiments identified a new sigma factor that directs galP1 transcription in vitro. DNase I protection experiments identified a binding site for this new holoenzyme immediately upstream of the galP1 transcription start site.
Brawner
M E
ME
Gene Expression Sciences, SmithKline Beecham Pharmaceuticals, King of Prussia, Pennsylvania 19406, USA.
Mattern
S G
SG
Babcock
M J
MJ
Westpheling
J
J
D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000235:genetics* / Q000378:metabolism; D005690:Galactose / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009876:Operon; D011401:Promoter Regions, Genetic; D012329:RNA, Bacterial / Q000032:analysis; D012808:Sigma Factor / Q000235:genetics; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics*; D014158:Transcription, Genetic
D012329:RNA, Bacterial; D012808:Sigma Factor; D012321:DNA-Directed RNA Polymerases; D005690:Galactose