Stopped in its tracks: The RNA polymerase molecular motor as a robust sensor of DNA damage

DNA repair is often a complex, multi-component, multi-step process; this makes detailed kinetic analysis of the different steps of repair a challenging task using standard biochemical methods. At the same time, single-molecule methods are well-suited for extracting kinetic information despite time-a...

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Bibliographic Details
Published in:DNA repair 2014-08, Vol.20, p.49-57
Main Authors: Howan, K., Monnet, J., Fan, J., Strick, T.R.
Format: Article
Language:English
Subjects:
Bacterial Proteins - chemistry
Bacterial Proteins - metabolism
DNA - chemistry
DNA - metabolism
DNA Damage
DNA Repair
DNA-Directed RNA Polymerases - chemistry
DNA-Directed RNA Polymerases - metabolism
Endodeoxyribonucleases - chemistry
Endodeoxyribonucleases - metabolism
Escherichia coli
Escherichia coli - chemistry
Escherichia coli - enzymology
Escherichia coli - genetics
Escherichia coli Proteins - chemistry
Escherichia coli Proteins - metabolism
Helicase
Magnetic trapping
Molecular Motor Proteins - chemistry
Molecular Motor Proteins - metabolism
Protein Binding
Repair
Single-molecule
Transcription
Transcription Factors - chemistry
Transcription Factors - metabolism
Transcription, Genetic
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Summary:DNA repair is often a complex, multi-component, multi-step process; this makes detailed kinetic analysis of the different steps of repair a challenging task using standard biochemical methods. At the same time, single-molecule methods are well-suited for extracting kinetic information despite time-averaging due to diffusion of biochemical components and stochasticity of chemical reaction steps. Here we discuss recent experiments using DNA nanomanipulation in a magnetic trap to study the initiation of transcription-coupled repair in a model bacterial system comprising the canonical Escherichia coli RNA polymerase and the Mfd translocase which specifically binds to it. These experiments provide kinetic insight into the reaction process, helping to explain how Mfd discriminates between transcribing RNAP and stalled RNAP. They also identify a reliably long-lived intermediate containing Mfd translocase and, potentially, RNA polymerase. This intermediate presumably serves as a platform for assembly of downstream repair components UvrAB(C).
ISSN:1568-7864
1568-7856
DOI:10.1016/j.dnarep.2014.02.018