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Updated: May 5, 2026
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Direct Restart of a Replication Fork Stalled by a Head-On RNA Polymerase
Published on: April 30, 2010
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Backtracking by single RNA polymerase molecules observed at near-base-pair resolution
Joshua W Shaevitz1, Elio A Abbondanzieri, Robert Landick
1Department of Physics, Stanford University, Stanford, California 94305, USA.
Nature
|November 25, 2003
Summary
Escherichia coli RNA polymerase (RNAP) proofreading involves backtracking and cleavage. This study observed single RNAP molecules backtracking and recovering, revealing factors influencing pause duration.
Area of Science:
- Molecular Biology
- Biophysics
Background:
- Escherichia coli RNA polymerase (RNAP) synthesizes RNA with high fidelity.
- A proposed proofreading mechanism involves backtracking and endonucleolytic cleavage.
Purpose of the Study:
- To observe and characterize the backtracking and recovery mechanism of single RNAP molecules.
- To investigate factors influencing RNAP pausing during transcription.
Main Methods:
- Utilized ultrastable optical trapping apparatus.
- Developed a novel two-bead assay for near-base-pair precision monitoring of transcription.
Main Results:
- Observed single RNAP molecules undergoing backtracking (approx. 5 bp) and recovery.
- Backtracking events were infrequent, associated with pauses from 20s to over 30 min.
- Inosine triphosphate increased backtracking pause frequency; GreA/GreB proteins decreased pause duration.
Conclusions:
- Directly observed the backtracking and recovery mechanism in single RNAP molecules.
- Identified factors affecting RNAP pausing and proofreading efficiency.
- Provides insights into the fidelity of RNA synthesis.
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