Single-molecule FRET unveils induced-fit mechanism for substrate selectivity in flap endonuclease 1
- King Abdullah University of Science and Technology, Saudi Arabia
- Georgia State University, United States
- Lawrence Berkeley National Laboratory, United States
Abstract
Human flap endonuclease 1 (FEN1) and related structure-specific 5'nucleases precisely identify and incise aberrant DNA structures during replication, repair and recombination to avoid genomic instability. Yet, it is unclear how the 5'nuclease mechanisms of DNA distortion and protein ordering robustly mediate efficient and accurate substrate recognition and catalytic selectivity. Here, single-molecule sub-millisecond and millisecond analyses of FEN1 reveal a protein-DNA induced-fit mechanism that efficiently verifies substrate and suppresses off-target cleavage. FEN1 sculpts DNA with diffusion-limited kinetics to test DNA substrate. This DNA distortion mutually 'locks' protein and DNA conformation and enables substrate verification with extreme precision. Strikingly, FEN1 never misses cleavage of its cognate substrate while blocking probable formation of catalytically competent interactions with noncognate substrates and fostering their pre-incision dissociation. These findings establish FEN1 has practically perfect precision and that separate control of induced-fit substrate recognition sets up the catalytic selectivity of the nuclease active site for genome stability.
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Funding
King Abdullah University of Science and Technology (2201 CRG3)
- Fahad Rashid
- Paul D Harris
- Manal S Zaher
- Mohamed A Sobhy
- Luay I Joudeh
- Hubert Piwonski
- John A Tainer
- Satoshi Habuchi
- Samir M Hamdan
National Science Foundation (MCB-1149521)
- Chunli Yan
- Ivaylo Ivanov
NIH Clinical Center (R01GM110387)
- Chunli Yan
- Ivaylo Ivanov
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2017, Rashid et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Single-molecule FRET unveils induced-fit mechanism for substrate selectivity in flap endonuclease 1
eLife 6:e21884.
https://doi.org/10.7554/eLife.21884
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