Single-molecule imaging of UvrA and UvrB recruitment to DNA lesions in living Escherichia coli
Nucleotide excision repair (NER) removes chemically diverse DNA lesions in all domains of life. In Escherichia coli, UvrA and UvrB initiate NER, although the mechanistic details of how this occurs in vivo remain to be established. Here we provide, using single-molecule fluorescence imaging, a compre...
| Main Authors: | , , , , , , |
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| Format: | Journal article |
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Nature Publishing Group
2016
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| _version_ | 1826284407820910592 |
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| author | Stracy, M Jaciuk, M Uphoff, S Kapanidis, A Nowotny, M Sherratt, D Zawadzki, P |
| author_facet | Stracy, M Jaciuk, M Uphoff, S Kapanidis, A Nowotny, M Sherratt, D Zawadzki, P |
| author_sort | Stracy, M |
| collection | OXFORD |
| description | Nucleotide excision repair (NER) removes chemically diverse DNA lesions in all domains of life. In Escherichia coli, UvrA and UvrB initiate NER, although the mechanistic details of how this occurs in vivo remain to be established. Here we provide, using single-molecule fluorescence imaging, a comprehensive characterization of the lesion search, recognition and verification process in living cells. We show that NER initiation involves a two-step mechanism in which UvrA scans the genome and locates DNA damage independently of UvrB. Then UvrA recruits UvrB from solution to the lesion. These steps are coordinated by ATP binding and hydrolysis in the ‘proximal’ and ‘distal’ UvrA ATP-binding sites. We show that initial UvrB-independent damage recognition by UvrA requires ATPase activity in the distal site only. Subsequent UvrB recruitment requires ATP hydrolysis in the proximal site. Finally, UvrA is dissociated from the lesion complex, allowing UvrB to orchestrate the downstream NER reactions. |
| first_indexed | 2024-03-07T01:13:23Z |
| format | Journal article |
| id | oxford-uuid:8dd0f729-31fb-4540-9a1b-021d41dc7144 |
| institution | University of Oxford |
| last_indexed | 2024-03-07T01:13:23Z |
| publishDate | 2016 |
| publisher | Nature Publishing Group |
| record_format | dspace |
| spelling | oxford-uuid:8dd0f729-31fb-4540-9a1b-021d41dc71442022-03-26T22:53:35ZSingle-molecule imaging of UvrA and UvrB recruitment to DNA lesions in living Escherichia coliJournal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:8dd0f729-31fb-4540-9a1b-021d41dc7144Symplectic Elements at OxfordNature Publishing Group2016Stracy, MJaciuk, MUphoff, SKapanidis, ANowotny, MSherratt, DZawadzki, PNucleotide excision repair (NER) removes chemically diverse DNA lesions in all domains of life. In Escherichia coli, UvrA and UvrB initiate NER, although the mechanistic details of how this occurs in vivo remain to be established. Here we provide, using single-molecule fluorescence imaging, a comprehensive characterization of the lesion search, recognition and verification process in living cells. We show that NER initiation involves a two-step mechanism in which UvrA scans the genome and locates DNA damage independently of UvrB. Then UvrA recruits UvrB from solution to the lesion. These steps are coordinated by ATP binding and hydrolysis in the ‘proximal’ and ‘distal’ UvrA ATP-binding sites. We show that initial UvrB-independent damage recognition by UvrA requires ATPase activity in the distal site only. Subsequent UvrB recruitment requires ATP hydrolysis in the proximal site. Finally, UvrA is dissociated from the lesion complex, allowing UvrB to orchestrate the downstream NER reactions. |
| spellingShingle | Stracy, M Jaciuk, M Uphoff, S Kapanidis, A Nowotny, M Sherratt, D Zawadzki, P Single-molecule imaging of UvrA and UvrB recruitment to DNA lesions in living Escherichia coli |
| title | Single-molecule imaging of UvrA and UvrB
recruitment to DNA lesions in living Escherichia coli |
| title_full | Single-molecule imaging of UvrA and UvrB
recruitment to DNA lesions in living Escherichia coli |
| title_fullStr | Single-molecule imaging of UvrA and UvrB
recruitment to DNA lesions in living Escherichia coli |
| title_full_unstemmed | Single-molecule imaging of UvrA and UvrB
recruitment to DNA lesions in living Escherichia coli |
| title_short | Single-molecule imaging of UvrA and UvrB
recruitment to DNA lesions in living Escherichia coli |
| title_sort | single molecule imaging of uvra and uvrb recruitment to dna lesions in living escherichia coli |
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