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Title: Single-molecule FRET unveils induced-fit mechanism for substrate selectivity in flap endonuclease 1

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.

Authors:
 [1];  [1];  [1];  [1];  [1];  [2]; ORCiD logo [1];  [3];  [2];  [4];  [1]; ORCiD logo [1]
  1. Division of Biological and Environmental Science and Engineering, King Abdullah University of Science and Technology, Thuwal, Saudi Arabia
  2. Department of Chemistry, Georgia State University, Atlanta, United States, Center for Diagnostics and Therapeutics, Georgia State University, Atlanta, United States
  3. Lawrence Berkeley National Laboratory, Berkeley, United States
  4. Lawrence Berkeley National Laboratory, Berkeley, United States, Department of Molecular and Cellular Oncology, University of Texas MD Anderson Cancer Center, Houston, United States
Publication Date:
Sponsoring Org.:
USDOE
OSTI Identifier:
1347470
Alternate Identifier(s):
OSTI ID: 1347471
Grant/Contract Number:  
AC02-05CH11231
Resource Type:
Published Article
Journal Name:
eLife
Additional Journal Information:
Journal Name: eLife Journal Volume: 6; Journal ID: ISSN 2050-084X
Publisher:
eLife Sciences Publications, Ltd.
Country of Publication:
United States
Language:
English

Citation Formats

Rashid, Fahad, Harris, Paul D., Zaher, Manal S., Sobhy, Mohamed A., Joudeh, Luay I., Yan, Chunli, Piwonski, Hubert, Tsutakawa, Susan E., Ivanov, Ivaylo, Tainer, John A., Habuchi, Satoshi, and Hamdan, Samir M. Single-molecule FRET unveils induced-fit mechanism for substrate selectivity in flap endonuclease 1. United States: N. p., 2017. Web. doi:10.7554/eLife.21884.
Rashid, Fahad, Harris, Paul D., Zaher, Manal S., Sobhy, Mohamed A., Joudeh, Luay I., Yan, Chunli, Piwonski, Hubert, Tsutakawa, Susan E., Ivanov, Ivaylo, Tainer, John A., Habuchi, Satoshi, & Hamdan, Samir M. Single-molecule FRET unveils induced-fit mechanism for substrate selectivity in flap endonuclease 1. United States. doi:10.7554/eLife.21884.
Rashid, Fahad, Harris, Paul D., Zaher, Manal S., Sobhy, Mohamed A., Joudeh, Luay I., Yan, Chunli, Piwonski, Hubert, Tsutakawa, Susan E., Ivanov, Ivaylo, Tainer, John A., Habuchi, Satoshi, and Hamdan, Samir M. Thu . "Single-molecule FRET unveils induced-fit mechanism for substrate selectivity in flap endonuclease 1". United States. doi:10.7554/eLife.21884.
@article{osti_1347470,
title = {Single-molecule FRET unveils induced-fit mechanism for substrate selectivity in flap endonuclease 1},
author = {Rashid, Fahad and Harris, Paul D. and Zaher, Manal S. and Sobhy, Mohamed A. and Joudeh, Luay I. and Yan, Chunli and Piwonski, Hubert and Tsutakawa, Susan E. and Ivanov, Ivaylo and Tainer, John A. and Habuchi, Satoshi and Hamdan, Samir M.},
abstractNote = {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.},
doi = {10.7554/eLife.21884},
journal = {eLife},
number = ,
volume = 6,
place = {United States},
year = {2017},
month = {2}
}

Journal Article:
Free Publicly Available Full Text
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DOI: 10.7554/eLife.21884

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Cited by: 11 works
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