Escherichia coli RNA polymerase binding to a DNA terminus prevents formation of a closed promoter complex.
A 302 bp DNA fragment and a 113 bp subfragment of the former, both containing the fd gene VIII promoter (P VIII), were found to exhibit temperature-dependent differential behaviour in RNA chain initiation from P VIII. At 37 degrees C no significant differences were observed, while at 17 degrees C chain initiation was strongly suppressed only with the 113 bp fragment. This phenomenon depended on the presence of the (blunt) DNA terminus upstream from P VIII (position -70). Footprinting revealed that at 17 degrees C RNA polymerase was bound to this DNA fragment in a different mode. Contacts were observed only upstream from position -25. On the contrary, at 37 degrees C only the promoter complex footprint was visible. These results indicate that at 17 degrees C formation of the non-initiating complex is more favourable than formation of the promoter complex (which is closed at 17 degrees C; Hofer, B., Müller, D. and Köster, H. (1985) Nucleic Acids Res. 13, 5995-6013) and that formation of both complexes is mutually exclusive. No footprints of RNA polymerase were observed at other DNA termini. This indicates a sequence-specificity for the interaction at the terminus of the 113 bp fragment. The footprint pattern, together with features of the DNA sequence, suggests that the contacts involved in this interaction are similar to those promoter contacts formed upstream from position -20 and that DNA without a -10 region can be specifically recognized by RNA polymerase.
Hofer
B
B
Institut für Organische Chemie und Biochemie der Universität Hamburg, F.R.G.
Müller
D
D
Köster
H
H
D001426:Bacterial Proteins / Q000378:metabolism*; D001435:Bacteriophages / Q000235:genetics; D004279:DNA, Viral / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D005814:Genes, Viral; D009710:Nucleotide Mapping; D011401:Promoter Regions, Genetic
D001426:Bacterial Proteins; D004279:DNA, Viral; D012321:DNA-Directed RNA Polymerases