This article is missing information about homologs including other Cas9s, other Class 2 Type II CRISPR systems.(September 2021) |
CRISPR-associated endonuclease Cas9 | |||||||
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Identifiers | |||||||
Organism | |||||||
Symbol | cas9 | ||||||
Alt. symbols | SpCas9 | ||||||
Entrez | 901176 | ||||||
PDB | 4OO8 | ||||||
RefSeq (mRNA) | NC_002737.2 | ||||||
RefSeq (Prot) | NP_269215.1 | ||||||
UniProt | Q99ZW2 | ||||||
Other data | |||||||
EC number | 3.1.-.- | ||||||
Chromosome | Genomic: 0.85 - 0.86 Mb | ||||||
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Cas9 | |
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Identifiers | |
Symbol | ? |
InterPro | IPR028629 |
Cas9 (CRISPR associated protein 9, formerly called Cas5, Csn1, or Csx12) is a 160 kilodalton protein which plays a vital role in the immunological defense of certain bacteria against DNA viruses and plasmids, and is heavily utilized in genetic engineering applications. Its main function is to cut DNA and thereby alter a cell's genome. The CRISPR-Cas9 genome editing technique was a significant contributor to the Nobel Prize in Chemistry in 2020 being awarded to Emmanuelle Charpentier and Jennifer Doudna.[2]
More technically, Cas9 is a dual RNA-guided DNA endonuclease enzyme associated with the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) adaptive immune system in Streptococcus pyogenes.[3][4][5] S. pyogenes utilizes CRISPR to memorize and Cas9 to later interrogate and cleave foreign DNA, such as invading bacteriophage DNA or plasmid DNA.[4][6][7][8] Cas9 performs this interrogation by unwinding foreign DNA and checking for sites complementary to the 20 nucleotide spacer region of the guide RNA (gRNA). If the DNA substrate is complementary to the guide RNA, Cas9 cleaves the invading DNA. In this sense, the CRISPR-Cas9 mechanism has a number of parallels with the RNA interference (RNAi) mechanism in eukaryotes.
Apart from its original function in bacterial immunity, the Cas9 protein has been heavily utilized as a genome engineering tool to induce site-directed double-strand breaks in DNA. These breaks can lead to gene inactivation or the introduction of heterologous genes through non-homologous end joining and homologous recombination respectively in many laboratory model organisms. Research on the development of various cas9 variants has been a promising way of overcoming the limitation of the CRISPR-Cas9 genome editing. Some examples include Cas9 nickase (Cas9n), a variant that induces single-stranded breaks (SSBs) or variants recognizing different PAM sequences.[9] Alongside zinc finger nucleases and transcription activator-like effector nuclease (TALEN) proteins, Cas9 is becoming a prominent tool in the field of genome editing.
Cas9 has gained traction in recent years because it can cleave nearly any sequence complementary to the guide RNA.[4] Because the target specificity of Cas9 stems from the guide RNA:DNA complementarity and not modifications to the protein itself (like TALENs and zinc fingers), engineering Cas9 to target new DNA is straightforward.[10] Versions of Cas9 that bind but do not cleave cognate DNA can be used to locate transcriptional activator or repressors to specific DNA sequences in order to control transcriptional activation and repression.[11][12] Native Cas9 requires a guide RNA composed of two disparate RNAs that associate – the CRISPR RNA (crRNA), and the trans-activating crRNA (tracrRNA).[3] Cas9 targeting has been simplified through the engineering of a chimeric single guide RNA (chiRNA). Scientists have suggested that Cas9-based gene drives may be capable of editing the genomes of entire populations of organisms.[13] In 2015, Cas9 was used to modify the genome of human embryos for the first time.[14]
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