Abstract
Transcription-coupled repair (TCR), a subpathway of nucleotide excision repair (NER), has been known to lead to more efficient repair than the global NER repair (GGR). Here we use magnetic trapping of single DNA molecules to study the interactions of TCR proteins with stalled RNA polymerase (RNAP). Single RNAP molecules are stalled on a DNA transcript using either a CTP-less cassette or a thymine-thymine dimer located on the transcribed strand. Stalled RNAP is displaced by the Mfd translocase, and a long-lived intermediate is formed. We characterize the interaction between UvrA/UvrB proteins and the long-lived intermediate formed upon Mfd displacement of RNAP. This interaction leads to formation of a pre-incision complex that is catalytically competent for DNA incision, and the activity of this complex is tested for in the magnetic trap upon addition of UvrC.
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