Transcription initiation at a consensus bacterial promoter proceeds via a ‘bind-unwind-load-and-lock’ mechanism
Transcription initiation starts with unwinding of promoter DNA by RNA polymerase (RNAP) to form a catalytically competent RNAP-promoter complex (RPo). Despite extensive study, the mechanism of promoter unwinding has remained unclear, in part due to the transient nature of intermediates on path to RP...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
| Published: |
eLife Sciences Publications Ltd
2021-10-01
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| Series: | eLife |
| Subjects: | |
| Online Access: | https://elifesciences.org/articles/70090 |
| _version_ | 1811253443121119232 |
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| author | Abhishek Mazumder Richard H Ebright Achillefs N Kapanidis |
| author_facet | Abhishek Mazumder Richard H Ebright Achillefs N Kapanidis |
| author_sort | Abhishek Mazumder |
| collection | DOAJ |
| description | Transcription initiation starts with unwinding of promoter DNA by RNA polymerase (RNAP) to form a catalytically competent RNAP-promoter complex (RPo). Despite extensive study, the mechanism of promoter unwinding has remained unclear, in part due to the transient nature of intermediates on path to RPo. Here, using single-molecule unwinding-induced fluorescence enhancement to monitor promoter unwinding, and single-molecule fluorescence resonance energy transfer to monitor RNAP clamp conformation, we analyse RPo formation at a consensus bacterial core promoter. We find that the RNAP clamp is closed during promoter binding, remains closed during promoter unwinding, and then closes further, locking the unwound DNA in the RNAP active-centre cleft. Our work defines a new, ‘bind-unwind-load-and-lock’, model for the series of conformational changes occurring during promoter unwinding at a consensus bacterial promoter and provides the tools needed to examine the process in other organisms and at other promoters. |
| first_indexed | 2024-04-12T16:50:59Z |
| format | Article |
| id | doaj.art-a30f5b4d3ff2433d975fec7b77a92d47 |
| institution | Directory Open Access Journal |
| issn | 2050-084X |
| language | English |
| last_indexed | 2024-04-12T16:50:59Z |
| publishDate | 2021-10-01 |
| publisher | eLife Sciences Publications Ltd |
| record_format | Article |
| series | eLife |
| spelling | doaj.art-a30f5b4d3ff2433d975fec7b77a92d472022-12-22T03:24:24ZengeLife Sciences Publications LtdeLife2050-084X2021-10-011010.7554/eLife.70090Transcription initiation at a consensus bacterial promoter proceeds via a ‘bind-unwind-load-and-lock’ mechanismAbhishek Mazumder0https://orcid.org/0000-0002-9339-6256Richard H Ebright1https://orcid.org/0000-0001-8915-7140Achillefs N Kapanidis2https://orcid.org/0000-0001-6699-136XBiological Physics Research Group, Clarendon Laboratory, Department of Physics, University of Oxford, Oxford, United KingdomWaksman Institute and Department of Chemistry, Rutgers University, Piscataway, United StatesBiological Physics Research Group, Clarendon Laboratory, Department of Physics, University of Oxford, Oxford, United KingdomTranscription initiation starts with unwinding of promoter DNA by RNA polymerase (RNAP) to form a catalytically competent RNAP-promoter complex (RPo). Despite extensive study, the mechanism of promoter unwinding has remained unclear, in part due to the transient nature of intermediates on path to RPo. Here, using single-molecule unwinding-induced fluorescence enhancement to monitor promoter unwinding, and single-molecule fluorescence resonance energy transfer to monitor RNAP clamp conformation, we analyse RPo formation at a consensus bacterial core promoter. We find that the RNAP clamp is closed during promoter binding, remains closed during promoter unwinding, and then closes further, locking the unwound DNA in the RNAP active-centre cleft. Our work defines a new, ‘bind-unwind-load-and-lock’, model for the series of conformational changes occurring during promoter unwinding at a consensus bacterial promoter and provides the tools needed to examine the process in other organisms and at other promoters.https://elifesciences.org/articles/70090RNA polymerasesmFRETtranscription initiation |
| spellingShingle | Abhishek Mazumder Richard H Ebright Achillefs N Kapanidis Transcription initiation at a consensus bacterial promoter proceeds via a ‘bind-unwind-load-and-lock’ mechanism eLife RNA polymerase smFRET transcription initiation |
| title | Transcription initiation at a consensus bacterial promoter proceeds via a ‘bind-unwind-load-and-lock’ mechanism |
| title_full | Transcription initiation at a consensus bacterial promoter proceeds via a ‘bind-unwind-load-and-lock’ mechanism |
| title_fullStr | Transcription initiation at a consensus bacterial promoter proceeds via a ‘bind-unwind-load-and-lock’ mechanism |
| title_full_unstemmed | Transcription initiation at a consensus bacterial promoter proceeds via a ‘bind-unwind-load-and-lock’ mechanism |
| title_short | Transcription initiation at a consensus bacterial promoter proceeds via a ‘bind-unwind-load-and-lock’ mechanism |
| title_sort | transcription initiation at a consensus bacterial promoter proceeds via a bind unwind load and lock mechanism |
| topic | RNA polymerase smFRET transcription initiation |
| url | https://elifesciences.org/articles/70090 |
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