PAM-flexible genome editing with an engineered chimeric Cas9.
Journal
Nature communications
ISSN: 2041-1723
Titre abrégé: Nat Commun
Pays: England
ID NLM: 101528555
Informations de publication
Date de publication:
04 10 2023
04 10 2023
Historique:
received:
24
02
2023
accepted:
21
09
2023
medline:
6
10
2023
pubmed:
5
10
2023
entrez:
4
10
2023
Statut:
epublish
Résumé
CRISPR enzymes require a defined protospacer adjacent motif (PAM) flanking a guide RNA-programmed target site, limiting their sequence accessibility for robust genome editing applications. In this study, we recombine the PAM-interacting domain of SpRY, a broad-targeting Cas9 possessing an NRN > NYN (R = A or G, Y = C or T) PAM preference, with the N-terminus of Sc + +, a Cas9 with simultaneously broad, efficient, and accurate NNG editing capabilities, to generate a chimeric enzyme with highly flexible PAM preference: SpRYc. We demonstrate that SpRYc leverages properties of both enzymes to specifically edit diverse PAMs and disease-related loci for potential therapeutic applications. In total, the approaches to generate SpRYc, coupled with its robust flexibility, highlight the power of integrative protein design for Cas9 engineering and motivate downstream editing applications that require precise genomic positioning.
Identifiants
pubmed: 37794046
doi: 10.1038/s41467-023-41829-y
pii: 10.1038/s41467-023-41829-y
pmc: PMC10550912
doi:
Substances chimiques
CRISPR-Associated Protein 9
EC 3.1.-
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
6175Subventions
Organisme : Howard Hughes Medical Institute
Pays : United States
Organisme : NHLBI NIH HHS
ID : P01 HL142494
Pays : United States
Organisme : NCI NIH HHS
ID : DP2 CA281401
Pays : United States
Commentaires et corrections
Type : UpdateOf
Informations de copyright
© 2023. Springer Nature Limited.
Références
Methods Enzymol. 2014;546:1-20
pubmed: 25398333
Nature. 2016 Jan 28;529(7587):490-5
pubmed: 26735016
Nat Biotechnol. 2015 Feb;33(2):187-197
pubmed: 25513782
Eur J Neurol. 2018 Jan;25(1):24-34
pubmed: 28817209
Nat Protoc. 2021 Mar;16(3):1511-1547
pubmed: 33547443
Science. 2020 Apr 17;368(6488):290-296
pubmed: 32217751
Nat Commun. 2019 Feb 4;10(1):560
pubmed: 30718489
Nature. 2018 Apr 5;556(7699):57-63
pubmed: 29512652
Nucleic Acids Res. 2018 Jul 2;46(W1):W296-W303
pubmed: 29788355
Nature. 2017 Oct 19;550(7676):407-410
pubmed: 28931002
Nucleic Acids Res. 2014 Jun;42(10):6726-41
pubmed: 24744240
Science. 2013 Feb 15;339(6121):819-23
pubmed: 23287718
BMC Genomics. 2017 May 15;18(1):379
pubmed: 28506212
Microbiology (Reading). 2009 Mar;155(Pt 3):733-740
pubmed: 19246744
Methods Mol Biol. 2019;1961:29-44
pubmed: 30912038
Nat Biotechnol. 2015 Dec;33(12):1293-1298
pubmed: 26524662
Nat Med. 2018 Aug;24(8):1216-1224
pubmed: 30082871
Mol Cell. 2016 Apr 7;62(1):137-47
pubmed: 27041224
CRISPR J. 2022 Feb;5(1):123-130
pubmed: 35119294
Nature. 2014 Mar 6;507(7490):62-7
pubmed: 24476820
Nat Biotechnol. 2018 Oct;36(9):843-846
pubmed: 29813047
Science. 2018 Sep 21;361(6408):1259-1262
pubmed: 30166441
Nature. 2015 Jul 23;523(7561):481-5
pubmed: 26098369
RNA Biol. 2013 May;10(5):891-9
pubmed: 23403393
Nat Methods. 2020 Jun;17(6):559
pubmed: 32499620
Cell Rep. 2018 Jan 9;22(2):359-371
pubmed: 29320733
Nature. 2017 Nov 23;551(7681):464-471
pubmed: 29160308
Acta Crystallogr D Biol Crystallogr. 2010 Apr;66(Pt 4):486-501
pubmed: 20383002
Mol Cell. 2019 Feb 21;73(4):714-726.e4
pubmed: 30581144
Prog Mol Biol Transl Sci. 2021;181:31-43
pubmed: 34127199
Nat Biotechnol. 2020 Jul;38(7):883-891
pubmed: 32433547
Nat Biotechnol. 2019 Mar;37(3):224-226
pubmed: 30809026
Nature. 2016 Apr 20;533(7603):420-4
pubmed: 27096365
Cell. 2019 Aug 8;178(4):887-900.e14
pubmed: 31398342
Science. 2020 Jul 31;369(6503):566-571
pubmed: 32732424
Mol Cell. 2016 Mar 17;61(6):895-902
pubmed: 26990992
Science. 2012 Aug 17;337(6096):816-21
pubmed: 22745249
Science. 2013 Feb 15;339(6121):823-6
pubmed: 23287722
Nat Commun. 2020 May 18;11(1):2474
pubmed: 32424114
Nat Biotechnol. 2020 Oct;38(10):1154-1158
pubmed: 32393822
Nat Commun. 2020 Nov 2;11(1):5512
pubmed: 33139742
Neuromolecular Med. 2014 Jun;16(2):231-64
pubmed: 24615633
Sci Adv. 2018 Oct 24;4(10):eaau0766
pubmed: 30397647
Elife. 2013 Jan 29;2:e00471
pubmed: 23386978
Mol Biol Cell. 2018 Nov 15;29(23):2809-2820
pubmed: 30256717