CRISPR (clustered regularly interspaced short palindromic repeat) is an adaptive immune system that provides protection against mobile genetic elements (viruses, transposable elements and conjugative plasmids). CRISPR clusters contain spacers, sequences complementary to antecedent mobile elements, and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA). In type II CRISPR systems correct processing of pre-crRNA requires a trans-encoded small RNA (tracrRNA), endogenous ribonuclease 3 (rnc) and this protein (By similarity). The tracrRNA serves as a guide for ribonuclease 3-aided processing of pre-crRNA. Subsequently Cas9/crRNA/tracrRNA endonucleolytically cleaves linear or circular dsDNA target complementary to the spacer yielding blunt ends; Cas9 is inactive in the absence of the 2 guide RNAs (gRNA). Cas9 recognizes a 3'-G-rich protospacer adjacent motif (PAM, GGG in this organism) in the CRISPR repeat sequences to help distinguish self versus nonself, as targets within the bacterial CRISPR locus do not have PAMs. PAM recognition is also required for catalytic activity. Complements the gRNA coprocessing defect in a cas9 deletion in S.pyogenes strain 370, and cuts target DNA in Cas9:gRNAs mixing experiments with S.mutans strain UA159. Endonuclease activity on target dsDNA requires Mg(2+). Monomer (By similarity). Binds crRNA and tracrRNA. Has 2 endonuclease domains. The discontinuous RuvC-like domain cleaves the target DNA noncomplementary to crRNA while the HNH nuclease domain cleaves the target DNA complementary to crRNA. The PAM-interacting domain (PI domain, approximately residues 1102-1388) recognizes the PAM motif; swapping the PI domain of this enzyme with that from S.pyogenes Cas9 (AC Q99ZW2) prevents cleavage of DNA with the endogenous PAM site but confers the ability to cleave DNA with the PAM site specific for S.pyogenes CRISPRs. Coexpression of Cas9 and both gRNAs in human cells has shown it is possible to use this system to target and modify a DNA sequence of interest in situ. Belongs to the CRISPR-associated protein Cas9 family. Subtype II-A subfamily.