Bei Gong
Molecular insights into DNA interference by CRISPR-associated nuclease-helicase Cas3
Gong, Bei; Shin, Minsang; Sun, Jiali; Jung, Che-Hun; Bolt, Edward L.; van der oost, John; Kim, Jeong-Sun
Authors
Minsang Shin
Jiali Sun
Che-Hun Jung
ED BOLT ED.BOLT@NOTTINGHAM.AC.UK
Professor of Molecular Biology
John van der oost
Jeong-Sun Kim
Abstract
Mobile genetic elements in bacteria are neutralized by a system based on clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) proteins. Type I CRISPR-Cas systems use a “Cascade” ribonucleoprotein complex to guide RNA specifically to complementary sequence in invader double-stranded DNA (dsDNA), a process called “interference.” After target recogni- tion by Cascade, formation of an R-loop triggers recruitment of a Cas3 nuclease-helicase, completing the interference process by destroying the invader dsDNA. To elucidate the molecular mecha- nism of CRISPR interference, we analyzed crystal structures of Cas3 from the bacterium Thermobaculum terrenum, with and without a bound ATP analog. The structures reveal a histidine-aspartate (HD)-type nuclease domain fused to superfamily-2 (SF2) helicase domains and a distinct C-terminal domain. Binding of ATP analog at the interface of the SF2 helicase RecA-like domains rearranges a motif V with implications for the enzyme mechanism. The HD- nucleolytic site contains two metal ions that are positioned at the end of a proposed nucleic acid-binding tunnel running through the SF2 helicase structure. This structural alignment suggests a mecha- nism for 3′ to 5′ nucleolytic processing of the displaced strand of invader DNA that is coordinated with ATP-dependent 3′ to 5′ trans- location of Cas3 along DNA. In agreement with biochemical studies, the presented Cas3 structures reveal important mechanistic details on the neutralization of genetic invaders by type I CRISPR-Cas systems.
Citation
Gong, B., Shin, M., Sun, J., Jung, C.-H., Bolt, E. L., van der oost, J., & Kim, J.-S. (2014). Molecular insights into DNA interference by CRISPR-associated nuclease-helicase Cas3. Proceedings of the National Academy of Sciences, 111(46), https://doi.org/10.1073/pnas.1410806111
Journal Article Type | Article |
---|---|
Acceptance Date | Sep 26, 2014 |
Online Publication Date | Nov 3, 2014 |
Publication Date | Nov 18, 2014 |
Deposit Date | Jul 19, 2016 |
Publicly Available Date | Jul 19, 2016 |
Journal | Proceedings of the National Academy of Sciences |
Print ISSN | 0027-8424 |
Electronic ISSN | 1091-6490 |
Publisher | National Academy of Sciences |
Peer Reviewed | Peer Reviewed |
Volume | 111 |
Issue | 46 |
DOI | https://doi.org/10.1073/pnas.1410806111 |
Keywords | Cas3, CRISPR, Cascade, Bacterial Immunity, Cas Proteins |
Public URL | https://nottingham-repository.worktribe.com/output/735239 |
Publisher URL | http://www.pnas.org/content/111/46/16359 |
Contract Date | Jul 19, 2016 |
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Copyright Statement
Copyright information regarding this work can be found at the following address: http://eprints.nottingham.ac.uk/end_user_agreement.pdf
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