PpCas9 from Pasteurella pneumotropica — a compact Type II-C Cas9 ortholog active in human cells
| Autor: | Maksim Kazalov, Mikhail Khodorkovskii, Anatolii Arseniev, Polina Selkova, Sergey Shmakov, T. O. Artamonova, Aleksandra Vasileva, Daria Artamonova, Olga Musharova, Ignatiy Goryanin, Tatyana Zyubko, Konstantin Severinov, Iana Fedorova, Georgii Pobegalov, Marina Abramova |
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| Rok vydání: | 2020 |
| Předmět: |
AcademicSubjects/SCI00010
Genetic Vectors Gene Expression Sequence alignment Computational biology Biology Genome 03 medical and health sciences 0302 clinical medicine Genome editing CRISPR-Associated Protein 9 Escherichia coli Genetics Humans Pasteurella pneumotropica CRISPR Clustered Regularly Interspaced Short Palindromic Repeats Amino Acid Sequence Guide RNA Cloning Molecular Rhodobacteraceae Gene 030304 developmental biology Gene Editing 0303 health sciences Nuclease Base Sequence Sequence Homology Amino Acid Nucleic Acid Enzymes Cas9 Recombinant Proteins HEK293 Cells biology.protein Nucleic Acid Conformation CRISPR-Cas Systems Sequence Alignment Genome Bacterial 030217 neurology & neurosurgery RNA Guide Kinetoplastida |
| Zdroj: | Nucleic Acids Research |
| ISSN: | 1362-4962 0305-1048 |
| DOI: | 10.1093/nar/gkaa998 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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