Dissociation of halted T7 RNA polymerase elongation complexes proceeds via a forward-translocation mechanism
UMass Chan Affiliations
Department of Psychiatry, Brudnick Neuropsychiatric Research InstituteGraduate School of Biomedical Sciences
Document Type
Journal ArticlePublication Date
2007-06-08Keywords
Base Sequence; Biological Transport; DNA, Viral; DNA-Directed RNA Polymerases; Enzyme Stability; Escherichia coli; Kinetics; Models, Genetic; Mutation; Promoter Regions (Genetics); Templates, Genetic; Transcription, Genetic; Viral ProteinsLife Sciences
Medicine and Health Sciences
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Show full item recordAbstract
A recent model for the mechanism of intrinsic transcription termination involves dissociation of the RNA from forward-translocated (hypertranslocated) states of the complex [Yarnell WS, Roberts JW (1999) Science, 284:611-615]. The current study demonstrates that halted elongation complexes of T7 RNA polymerase in the absence of termination signals can also dissociate via a forward-translocation mechanism. Shortening of the downstream DNA or the introduction of a stretch of mismatched DNA immediately downstream of the halt site reduces a barrier to forward translocation and correspondingly reduces the lifetime of halted complexes. Conversely, introduction of a cross-link downstream of the halt site increases the same barrier and leads to an increase in complex lifetime. Introduction of a mismatch within the bubble reduces a driving force for forward translocation and correspondingly increases the lifetime of the complex, but only for mismatches at the upstream edge of the bubble, as predicted by the model. Mismatching only the two most upstream of the eight bases in the bubble provides a maximal increase in complex stability, suggesting that dissociation occurs primarily from early forward-translocated states. Finally, addition in trans of an oligonucleotide complementary to the nascent RNA just beyond the hybrid complements the loss of driving force derived from placement of a mismatch within the bubble, confirming the expected additivity of effects. Thus, forward translocation is likely a general mechanism for dissociation of elongation complexes, both in the presence and absence of intrinsic termination signals.Source
Proc Natl Acad Sci U S A. 2007 Jun 19;104(25):10352-7. Epub 2007 Jun 6. Link to article on publisher's siteDOI
10.1073/pnas.0606306104Permanent Link to this Item
http://hdl.handle.net/20.500.14038/33772PubMed ID
17553968Related Resources
Link to Article in PubMedae974a485f413a2113503eed53cd6c53
10.1073/pnas.0606306104