Backtracking by single RNA polymerase molecules observed at near-base-pair resolution
| Autor: | Robert Landick, Steven M. Block, Joshua W. Shaevitz, Elio A. Abbondanzieri |
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| Rok vydání: | 2003 |
| Předmět: |
Transcription
Genetic Base pair Biology Cleavage (embryo) Article Substrate Specificity chemistry.chemical_compound Transcription (biology) RNA polymerase Escherichia coli Binding site Base Pairing Binding Sites Multidisciplinary Escherichia coli Proteins RNA DNA DNA-Directed RNA Polymerases Templates Genetic RNA Bacterial chemistry Biochemistry Mutagenesis Biophysics Proofreading Transcriptional Elongation Factors Transcription Factors |
| Zdroj: | Nature. 426:684-687 |
| ISSN: | 1476-4687 0028-0836 |
| Popis: | Escherichia coli RNA polymerase (RNAP) synthesizes RNA with remarkable fidelity in vivo. Its low error rate may be achieved by means of a 'proofreading' mechanism comprised of two sequential events. The first event (backtracking) involves a transcriptionally upstream motion of RNAP through several base pairs, which carries the 3' end of the nascent RNA transcript away from the enzyme active site. The second event (endonucleolytic cleavage) occurs after a variable delay and results in the scission and release of the most recently incorporated ribonucleotides, freeing up the active site. Here, by combining ultrastable optical trapping apparatus with a novel two-bead assay to monitor transcriptional elongation with near-base-pair precision, we observed backtracking and recovery by single molecules of RNAP. Backtracking events ( approximately 5 bp) occurred infrequently at locations throughout the DNA template and were associated with pauses lasting 20 s to30 min. Inosine triphosphate increased the frequency of backtracking pauses, whereas the accessory proteins GreA and GreB, which stimulate the cleavage of nascent RNA, decreased the duration of such pauses. |
| Databáze: | OpenAIRE |
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