Probing the mechanics of the complete DNA transcription cycle in real-time using optical tweezers

Christoph G Baumann, Stephen J Cross

Research output: Contribution to journalArticle (Academic Journal)

2 Citations (Scopus)

Abstract

RNA polymerase (RNAP) is a DNA-dependent motor protein that links ribonucleotide polymerization to force generation and DNA translocation through its active site, i.e., mechanical work. Single-molecule studies using optical tweezers have allowed researchers to probe the load-dependent ribonucleotide incorporation rate and processivity of both single-subunit viral and multisubunit prokaryotic and eukaryotic RNAPs engaged in transcription elongation. A single-molecule method is described here, which allows the complete transcription cycle (i.e., promoter binding, initiation, elongation and termination) to be followed in real-time using dual-trap optical tweezers and a unique "three-bead" geometry. This single-molecule transcription assay can be used to probe the mechanics of both stationary and moving RNAP-DNA complexes engaged in different stages of transcription.

Original languageEnglish
Pages (from-to)175-91
Number of pages17
JournalMethods in Molecular Biology
Volume778
DOIs
Publication statusPublished - 2011

Keywords

  • DNA-Directed RNA Polymerases
  • Molecular Motor Proteins
  • Optical Tweezers
  • Transcription, Genetic
  • Viral Proteins

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