PpCas9 from Pasteurella pneumotropica - a compact Type II-C Cas9 ortholog active in human cells.
Amino Acid Sequence
Base Sequence
CRISPR-Associated Protein 9
/ chemistry
CRISPR-Cas Systems
Cloning, Molecular
Clustered Regularly Interspaced Short Palindromic Repeats
Escherichia coli
/ genetics
Gene Editing
/ methods
Gene Expression
Genetic Vectors
/ chemistry
Genome, Bacterial
HEK293 Cells
Humans
Nucleic Acid Conformation
Pasteurella pneumotropica
/ enzymology
RNA, Guide, Kinetoplastida
/ chemistry
Recombinant Proteins
/ chemistry
Rhodobacteraceae
/ enzymology
Sequence Alignment
Sequence Homology, Amino Acid
Journal
Nucleic acids research
ISSN: 1362-4962
Titre abrégé: Nucleic Acids Res
Pays: England
ID NLM: 0411011
Informations de publication
Date de publication:
02 12 2020
02 12 2020
Historique:
accepted:
16
10
2020
revised:
11
10
2020
received:
04
05
2020
pubmed:
6
11
2020
medline:
30
12
2020
entrez:
5
11
2020
Statut:
ppublish
Résumé
CRISPR-Cas defense systems opened up the field of genome editing due to the ease with which effector Cas nucleases can be programmed with guide RNAs to access desirable genomic sites. Type II-A SpCas9 from Streptococcus pyogenes was the first Cas9 nuclease used for genome editing and it remains the most popular enzyme of its class. Nevertheless, SpCas9 has some drawbacks including a relatively large size and restriction to targets flanked by an 'NGG' PAM sequence. The more compact Type II-C Cas9 orthologs can help to overcome the size limitation of SpCas9. Yet, only a few Type II-C nucleases were fully characterized to date. Here, we characterized two Cas9 II-C orthologs, DfCas9 from Defluviimonas sp.20V17 and PpCas9 from Pasteurella pneumotropica. Both DfCas9 and PpCas9 cleave DNA in vitro and have novel PAM requirements. Unlike DfCas9, the PpCas9 nuclease is active in human cells. This small nuclease requires an 'NNNNRTT' PAM orthogonal to that of SpCas9 and thus potentially can broaden the range of Cas9 applications in biomedicine and biotechnology.
Identifiants
pubmed: 33152077
pii: 5957178
doi: 10.1093/nar/gkaa998
pmc: PMC7708072
doi:
Substances chimiques
RNA, Guide
0
Recombinant Proteins
0
CRISPR-Associated Protein 9
EC 3.1.-
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
12297-12309Informations de copyright
© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.
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