Nucleic Acids Research, 2002, Vol. 30, No. 4 886-893
© 2002 Oxford University Press
Interactions of regulated and deregulated forms of the 
54 holoenzyme with heteroduplex promoter DNA
Department of Biological Sciences, Imperial College of Science, Technology and Medicine, Sir Alexander Fleming Building, Imperial College Road, London SW7 2AZ, UK
The bacterial 
54 RNA polymerase holoenzyme binds to promoters as a stable closed complex that is silent for transcription unless acted upon by an enhancer-bound activator protein. Using DNA binding and transcription assays the ability of the enhancer-dependent 54 holoenzyme to interact with promoter DNA containing various regions of heteroduplex from –12 to –1 was assessed. Different DNA regions important for stabilising 54 holoenzyme–promoter interactions, destabilising binding, limiting template utilisation in activator-dependent transcription and for stable binding of a deregulated form of the holoenzyme lacking 54 Region I were identified. It appears that homoduplex structures are required for early events in 54 holoenzyme promoter binding and that disruption of a repressive fork junction structure only modestly deregulates transcription. DNA opening from –5 to –1 appears important for stable engagement of the holoenzyme following activation. The regulatory Region I of 54 was shown to be involved in interactions with the sequences in the –5 to –1 area.
* To whom correspondence should be addressed. Tel: +44 207 594 5442; Fax: +44 207 594 5419; Email: m.buck{at}ic.ac.uk
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