Protospacer Adjacent Motif-Induced Allostery Activates CRISPR-Cas9
Abstract
CRISPR-Cas9 is a genome editing technology with major impact in life sciences. In this system, the endonuclease Cas9 generates double strand breaks in DNA upon RNA-guided recognition of a complementary DNA sequence, which strictly requires the presence of a protospacer adjacent motif (PAM) next to the target site. Although PAM recognition is essential for cleavage, it is unknown whether and how PAM binding activates Cas9 for DNA cleavage at spatially distant sites. Here, we find evidence of a PAM-induced allosteric mechanism revealed by microsecond molecular dynamics simulations. PAM acts as an allosteric effector and triggers the interdependent conformational dynamics of the Cas9 catalytic domains (HNH and RuvC), responsible for concerted cleavage of the two DNA strands. As a result, targeting such an allosteric mechanism should enable control of CRISPR-Cas9 functionality.
- Authors:
-
[1];
[1];
[2];
[4];
[2];
- Univ. of California San Diego, La Jolla, CA (United States)
- Yale Univ., New Haven, CT (United States)
- Univ. of Zurich, Zurich (Switzerland)
- Univ. Claude Bernard Lyon, Lyon (France)
- Publication Date:
- Research Org.:
- Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States). National Energy Research Scientific Computing Center (NERSC)
- Sponsoring Org.:
- USDOE
- OSTI Identifier:
- 1489021
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Journal of the American Chemical Society
- Additional Journal Information:
- Journal Volume: 139; Journal Issue: 45; Journal ID: ISSN 0002-7863
- Publisher:
- American Chemical Society (ACS)
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 60 APPLIED LIFE SCIENCES
Citation Formats
Palermo, Giulia, Ricci, Clarisse G., Fernando, Amendra, Basak, Rajshekhar, Jinek, Martin, Rivalta, Ivan, Batista, Victor S., and McCammon, J. Andrew. Protospacer Adjacent Motif-Induced Allostery Activates CRISPR-Cas9. United States: N. p., 2017.
Web. doi:10.1021/jacs.7b05313.
Palermo, Giulia, Ricci, Clarisse G., Fernando, Amendra, Basak, Rajshekhar, Jinek, Martin, Rivalta, Ivan, Batista, Victor S., & McCammon, J. Andrew. Protospacer Adjacent Motif-Induced Allostery Activates CRISPR-Cas9. United States. https://doi.org/10.1021/jacs.7b05313
Palermo, Giulia, Ricci, Clarisse G., Fernando, Amendra, Basak, Rajshekhar, Jinek, Martin, Rivalta, Ivan, Batista, Victor S., and McCammon, J. Andrew. Wed .
"Protospacer Adjacent Motif-Induced Allostery Activates CRISPR-Cas9". United States. https://doi.org/10.1021/jacs.7b05313. https://www.osti.gov/servlets/purl/1489021.
@article{osti_1489021,
title = {Protospacer Adjacent Motif-Induced Allostery Activates CRISPR-Cas9},
author = {Palermo, Giulia and Ricci, Clarisse G. and Fernando, Amendra and Basak, Rajshekhar and Jinek, Martin and Rivalta, Ivan and Batista, Victor S. and McCammon, J. Andrew},
abstractNote = {CRISPR-Cas9 is a genome editing technology with major impact in life sciences. In this system, the endonuclease Cas9 generates double strand breaks in DNA upon RNA-guided recognition of a complementary DNA sequence, which strictly requires the presence of a protospacer adjacent motif (PAM) next to the target site. Although PAM recognition is essential for cleavage, it is unknown whether and how PAM binding activates Cas9 for DNA cleavage at spatially distant sites. Here, we find evidence of a PAM-induced allosteric mechanism revealed by microsecond molecular dynamics simulations. PAM acts as an allosteric effector and triggers the interdependent conformational dynamics of the Cas9 catalytic domains (HNH and RuvC), responsible for concerted cleavage of the two DNA strands. As a result, targeting such an allosteric mechanism should enable control of CRISPR-Cas9 functionality.},
doi = {10.1021/jacs.7b05313},
journal = {Journal of the American Chemical Society},
number = 45,
volume = 139,
place = {United States},
year = {Wed Aug 02 00:00:00 EDT 2017},
month = {Wed Aug 02 00:00:00 EDT 2017}
}
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Cited by: 73 works
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Figures / Tables:
Figure 1.: Architecture of the S. pyogenes CRISPR-Cas9 bound to a guide RNA and a target DNA (4UN3.pdb), with close-up view of the PAM recognition region. Two stars highlight the cleavage sites.
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Works referenced in this record:
The new frontier of genome engineering with CRISPR-Cas9
journal, November 2014
- Doudna, Jennifer A.; Charpentier, Emmanuelle
- Science, Vol. 346, Issue 6213
A Programmable Dual-RNA-Guided DNA Endonuclease in Adaptive Bacterial Immunity
journal, June 2012
- Jinek, M.; Chylinski, K.; Fonfara, I.
- Science, Vol. 337, Issue 6096, p. 816-821
DNA interrogation by the CRISPR RNA-guided endonuclease Cas9
journal, January 2014
- Sternberg, Samuel H.; Redding, Sy; Jinek, Martin
- Nature, Vol. 507, Issue 7490
CRISPR–Cas9 Structures and Mechanisms
journal, May 2017
- Jiang, Fuguo; Doudna, Jennifer A.
- Annual Review of Biophysics, Vol. 46, Issue 1
Structural basis of PAM-dependent target DNA recognition by the Cas9 endonuclease
journal, July 2014
- Anders, Carolin; Niewoehner, Ole; Duerst, Alessia
- Nature, Vol. 513, Issue 7519
Conformational control of DNA target cleavage by CRISPR–Cas9
journal, October 2015
- Sternberg, Samuel H.; LaFrance, Benjamin; Kaplan, Matias
- Nature, Vol. 527, Issue 7576
Crystal Structure of Cas9 in Complex with Guide RNA and Target DNA
journal, February 2014
- Nishimasu, Hiroshi; Ran, F. Ann; Hsu, Patrick D.
- Cell, Vol. 156, Issue 5
Catalytic Metal Ions and Enzymatic Processing of DNA and RNA
journal, January 2015
- Palermo, Giulia; Cavalli, Andrea; Klein, Michael L.
- Accounts of Chemical Research, Vol. 48, Issue 2
CRISPR-Cas9 conformational activation as elucidated from enhanced molecular simulations
journal, June 2017
- Palermo, Giulia; Miao, Yinglong; Walker, Ross C.
- Proceedings of the National Academy of Sciences, Vol. 114, Issue 28
Striking Plasticity of CRISPR-Cas9 and Key Role of Non-target DNA, as Revealed by Molecular Simulations
journal, September 2016
- Palermo, Giulia; Miao, Yinglong; Walker, Ross C.
- ACS Central Science, Vol. 2, Issue 10
Generalized correlation for biomolecular dynamics
journal, December 2005
- Lange, Oliver F.; Grubmüller, Helmut
- Proteins: Structure, Function, and Bioinformatics, Vol. 62, Issue 4
Protein Allostery and Conformational Dynamics
journal, February 2016
- Guo, Jingjing; Zhou, Huan-Xiang
- Chemical Reviews, Vol. 116, Issue 11
Allosteric cross-talk in chromatin can mediate drug-drug synergy
journal, March 2017
- Adhireksan, Zenita; Palermo, Giulia; Riedel, Tina
- Nature Communications, Vol. 8, Issue 1
Dynamical networks in tRNA:protein complexes
journal, April 2009
- Sethi, A.; Eargle, J.; Black, A. A.
- Proceedings of the National Academy of Sciences, Vol. 106, Issue 16
Allosteric pathways in imidazole glycerol phosphate synthase
journal, May 2012
- Rivalta, I.; Sultan, M. M.; Lee, N. -S.
- Proceedings of the National Academy of Sciences, Vol. 109, Issue 22
Allosteric Pathways in the PPARγ-RXRα nuclear receptor complex
journal, January 2016
- Ricci, Clarisse G.; Silveira, Rodrigo L.; Rivalta, Ivan
- Scientific Reports, Vol. 6, Issue 1
Structures of a CRISPR-Cas9 R-loop complex primed for DNA cleavage
journal, January 2016
- Jiang, F.; Taylor, D. W.; Chen, J. S.
- Science, Vol. 351, Issue 6275
Rationally engineered Cas9 nucleases with improved specificity
journal, December 2015
- Slaymaker, I. M.; Gao, L.; Zetsche, B.
- Science, Vol. 351, Issue 6268
Engineered CRISPR-Cas9 nucleases with altered PAM specificities
journal, June 2015
- Kleinstiver, Benjamin P.; Prew, Michelle S.; Tsai, Shengdar Q.
- Nature, Vol. 523, Issue 7561
Electrostatics of nanosystems: Application to microtubules and the ribosome
journal, August 2001
- Baker, N. A.; Sept, D.; Joseph, S.
- Proceedings of the National Academy of Sciences, Vol. 98, Issue 18, p. 10037-10041
Generalized correlation for biomolecular dynamics
journal, December 2005
- Lange, Oliver F.; Grubmüller, Helmut
- Proteins: Structure, Function, and Bioinformatics, Vol. 62, Issue 4
Improvement of Soybean; A Way Forward Transition from Genetic Engineering to New Plant Breeding Technologies
journal, February 2022
- Rahman, Saleem Ur; McCoy, Evan; Raza, Ghulam
- Molecular Biotechnology, Vol. 65, Issue 2
DNA Interrogation by the CRISPR RNA-Guided Endonuclease Cas9
journal, January 2014
- Sternberg, Samuel H.; Redding, Sy; Jinek, Martin
- Biophysical Journal, Vol. 106, Issue 2
Protein Allostery and Conformational Dynamics
journal, February 2016
- Guo, Jingjing; Zhou, Huan-Xiang
- Chemical Reviews, Vol. 116, Issue 11
Catalytic Metal Ions and Enzymatic Processing of DNA and RNA
journal, January 2015
- Palermo, Giulia; Cavalli, Andrea; Klein, Michael L.
- Accounts of Chemical Research, Vol. 48, Issue 2
Engineered CRISPR-Cas9 nucleases with altered PAM specificities
journal, June 2015
- Kleinstiver, Benjamin P.; Prew, Michelle S.; Tsai, Shengdar Q.
- Nature, Vol. 523, Issue 7561
Conformational control of DNA target cleavage by CRISPR–Cas9
journal, October 2015
- Sternberg, Samuel H.; LaFrance, Benjamin; Kaplan, Matias
- Nature, Vol. 527, Issue 7576
Allosteric cross-talk in chromatin can mediate drug-drug synergy
journal, March 2017
- Adhireksan, Zenita; Palermo, Giulia; Riedel, Tina
- Nature Communications, Vol. 8, Issue 1
Dynamical networks in tRNA:protein complexes
journal, April 2009
- Sethi, A.; Eargle, J.; Black, A. A.
- Proceedings of the National Academy of Sciences, Vol. 106, Issue 16
Allosteric pathways in imidazole glycerol phosphate synthase
journal, May 2012
- Rivalta, I.; Sultan, M. M.; Lee, N. -S.
- Proceedings of the National Academy of Sciences, Vol. 109, Issue 22
Rationally engineered Cas9 nucleases with improved specificity
journal, December 2015
- Slaymaker, I. M.; Gao, L.; Zetsche, B.
- Science, Vol. 351, Issue 6268
Structures of a CRISPR-Cas9 R-loop complex primed for DNA cleavage
journal, January 2016
- Jiang, F.; Taylor, D. W.; Chen, J. S.
- Science, Vol. 351, Issue 6275
Works referencing / citing this record:
Mechanism of allosteric activation of human mRNA cap methyltransferase (RNMT) by RAM: insights from accelerated molecular dynamics simulations
journal, July 2019
- Bueren-Calabuig, Juan A.; G. Bage, Marcus; Cowling, Victoria H.
- Nucleic Acids Research
Dynamics changes of CRISPR-Cas9 systems induced by high fidelity mutations
journal, January 2018
- Zheng, Liangzhen; Shi, Jiahai; Mu, Yuguang
- Physical Chemistry Chemical Physics, Vol. 20, Issue 43
Eigenvector centrality for characterization of protein allosteric pathways
journal, December 2018
- Negre, Christian F. A.; Morzan, Uriel N.; Hendrickson, Heidi P.
- Proceedings of the National Academy of Sciences, Vol. 115, Issue 52
1H, 13C, 15N backbone and side chain resonance assignment of the HNH nuclease from Streptococcus pyogenes CRISPR-Cas9
journal, August 2019
- Belato, Helen B.; East, Kyle W.; Lisi, George P.
- Biomolecular NMR Assignments, Vol. 13, Issue 2
Future of human mitochondrial DNA editing technologies
journal, May 2018
- Verechshagina, N.; Nikitchina, N.; Yamada, Y.
- Mitochondrial DNA Part A, Vol. 30, Issue 2
Key role of the REC lobe during CRISPR–Cas9 activation by ‘sensing’, ‘regulating’, and ‘locking’ the catalytic HNH domain
journal, January 2018
- Palermo, Giulia; Chen, Janice S.; Ricci, Clarisse G.
- Quarterly Reviews of Biophysics, Vol. 51
All-atom simulations disentangle the functional dynamics underlying gene maturation in the intron lariat spliceosome
journal, June 2018
- Casalino, Lorenzo; Palermo, Giulia; Spinello, Angelo
- Proceedings of the National Academy of Sciences, Vol. 115, Issue 26
Liposome-based transfection enhances RNAi and CRISPR-mediated mutagenesis in non-model nematode systems
journal, January 2019
- Adams, Sally; Pathak, Prachi; Shao, Hongguang
- Scientific Reports, Vol. 9, Issue 1
NMR and computational methods for molecular resolution of allosteric pathways in enzyme complexes
journal, December 2019
- East, Kyle W.; Skeens, Erin; Cui, Jennifer Y.
- Biophysical Reviews, Vol. 12, Issue 1
PAM identification by CRISPR-Cas effector complexes: diversified mechanisms and structures
journal, September 2018
- Gleditzsch, Daniel; Pausch, Patrick; Müller-Esparza, Hanna
- RNA Biology, Vol. 16, Issue 4
Allostery in Its Many Disguises: From Theory to Applications
journal, April 2019
- Wodak, Shoshana J.; Paci, Emanuele; Dokholyan, Nikolay V.
- Structure, Vol. 27, Issue 4
Key role of the REC lobe during CRISPR–Cas9 activation by ‘sensing’, ‘regulating’, and ‘locking’ the catalytic HNH domain
journal, January 2018
- Palermo, Giulia; Chen, Janice S.; Ricci, Clarisse G.
- Quarterly Reviews of Biophysics, Vol. 51
Disclosing the Impact of Carcinogenic SF3b Mutations on Pre-mRNA Recognition Via All-Atom Simulations
journal, October 2019
- Borišek, Jure; Saltalamacchia, Andrea; Gallì, Anna
- Biomolecules, Vol. 9, Issue 10
Figures / Tables found in this record:
Figures/Tables have been extracted from DOE-funded journal article accepted manuscripts.