CS12A_FRATN
ID CS12A_FRATN Reviewed; 1300 AA.
AC A0Q7Q2;
DT 09-DEC-2015, integrated into UniProtKB/Swiss-Prot.
DT 09-JAN-2007, sequence version 1.
DT 03-AUG-2022, entry version 68.
DE RecName: Full=CRISPR-associated endonuclease Cas12a {ECO:0000303|PubMed:28111461};
DE EC=3.1.21.1 {ECO:0000269|PubMed:28431230};
DE EC=4.6.1.22 {ECO:0000269|PubMed:28431230};
DE AltName: Full=CRISPR-associated endonuclease Cpf1 {ECO:0000303|PubMed:26422227};
DE AltName: Full=FnCas12a {ECO:0000303|PubMed:28431230};
DE AltName: Full=FnCpf1 {ECO:0000303|PubMed:26422227};
GN Name=cas12a {ECO:0000303|PubMed:28111461};
GN Synonyms=cpf1 {ECO:0000303|PubMed:26422227}; OrderedLocusNames=FTN_1397;
OS Francisella tularensis subsp. novicida (strain U112).
OC Bacteria; Proteobacteria; Gammaproteobacteria; Thiotrichales;
OC Francisellaceae; Francisella.
OX NCBI_TaxID=401614;
RN [1]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=U112;
RX PubMed=17550600; DOI=10.1186/gb-2007-8-6-r102;
RA Rohmer L., Fong C., Abmayr S., Wasnick M., Larson Freeman T.J., Radey M.,
RA Guina T., Svensson K., Hayden H.S., Jacobs M., Gallagher L.A., Manoil C.,
RA Ernst R.K., Drees B., Buckley D., Haugen E., Bovee D., Zhou Y., Chang J.,
RA Levy R., Lim R., Gillett W., Guenthener D., Kang A., Shaffer S.A.,
RA Taylor G., Chen J., Gallis B., D'Argenio D.A., Forsman M., Olson M.V.,
RA Goodlett D.R., Kaul R., Miller S.I., Brittnacher M.J.;
RT "Comparison of Francisella tularensis genomes reveals evolutionary events
RT associated with the emergence of human pathogenic strains.";
RL Genome Biol. 8:R102.1-R102.16(2007).
RN [2]
RP FUNCTION IN PLASMID RESISTANCE, FUNCTION IN CRRNA FORMATION, FUNCTION AS AN
RP ENDONUCLEASE, POSSIBLE ACTIVE SITE, COFACTOR, POSSIBLE SUBUNIT, AND
RP MUTAGENESIS OF ASP-917; GLU-1006 AND ASP-1255.
RC STRAIN=U112;
RX PubMed=26422227; DOI=10.1016/j.cell.2015.09.038;
RA Zetsche B., Gootenberg J.S., Abudayyeh O.O., Slaymaker I.M., Makarova K.S.,
RA Essletzbichler P., Volz S.E., Joung J., van der Oost J., Regev A.,
RA Koonin E.V., Zhang F.;
RT "Cpf1 is a single RNA-guided endonuclease of a class 2 CRISPR-Cas system.";
RL Cell 163:759-771(2015).
RN [3]
RP DISCUSSION OF SEQUENCE.
RX PubMed=26593719; DOI=10.1016/j.molcel.2015.10.008;
RA Shmakov S., Abudayyeh O.O., Makarova K.S., Wolf Y.I., Gootenberg J.S.,
RA Semenova E., Minakhin L., Joung J., Konermann S., Severinov K., Zhang F.,
RA Koonin E.V.;
RT "Discovery and functional characterization of diverse class 2 CRISPR-Cas
RT systems.";
RL Mol. Cell 60:385-397(2015).
RN [4]
RP FUNCTION IN CRRNA PROCESSING, FUNCTION AS AN ENDORIBONUCLEASE, FUNCTION AS
RP AN ENDONUCLEASE, COFACTOR, SUBUNIT, MUTAGENESIS OF HIS-843; LYS-852;
RP LYS-869; PHE-873; ASP-917; GLU-920; HIS-922; TYR-925; GLU-1006; TYR-1024;
RP GLU-1028; ASP-1227 AND ASP-1255, DNA-BINDING, AND RNA-BINDING.
RC STRAIN=U112;
RX PubMed=27096362; DOI=10.1038/nature17945;
RA Fonfara I., Richter H., Bratovic M., Le Rhun A., Charpentier E.;
RT "The CRISPR-associated DNA-cleaving enzyme Cpf1 also processes precursor
RT CRISPR RNA.";
RL Nature 532:517-521(2016).
RN [5]
RP BIOTECHNOLOGY.
RX PubMed=27905529; DOI=10.1038/srep38169;
RA Endo A., Masafumi M., Kaya H., Toki S.;
RT "Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from
RT Francisella novicida.";
RL Sci. Rep. 6:38169-38169(2016).
RN [6]
RP BIOTECHNOLOGY.
RX PubMed=27272384; DOI=10.1038/nbt.3609;
RA Kim D., Kim J., Hur J.K., Been K.W., Yoon S.H., Kim J.S.;
RT "Genome-wide analysis reveals specificities of Cpf1 endonucleases in human
RT cells.";
RL Nat. Biotechnol. 34:863-868(2016).
RN [7]
RP NOMENCLATURE.
RX PubMed=28111461; DOI=10.1038/nrmicro.2016.184;
RA Shmakov S., Smargon A., Scott D., Cox D., Pyzocha N., Yan W.,
RA Abudayyeh O.O., Gootenberg J.S., Makarova K.S., Wolf Y.I., Severinov K.,
RA Zhang F., Koonin E.V.;
RT "Diversity and evolution of class 2 CRISPR-Cas systems.";
RL Nat. Rev. Microbiol. 15:169-182(2017).
RN [8] {ECO:0007744|PDB:5NFV, ECO:0007744|PDB:5NG6}
RP X-RAY CRYSTALLOGRAPHY (2.50 ANGSTROMS) OF 2-1300 IN COMPLEX WITH GUIDE RNA
RP WITH AND WITHOUT TARGET DNA, FUNCTION IN CRRNA PROCESSING, FUNCTION AS AN
RP ENDONUCLEASE, ACTIVE SITE, CATALYTIC ACTIVITY, REACTION MECHANISM,
RP COFACTOR, SUBUNIT, DOMAIN, MUTAGENESIS OF ARG-692; 689-THR--SER-702;
RP GLN-704; ASP-917; GLU-1006; ARG-1218 AND ASP-1255, DNA-BINDING, AND
RP RNA-BINDING.
RC STRAIN=U112;
RX PubMed=28431230; DOI=10.1016/j.molcel.2017.03.016;
RA Swarts D.C., van der Oost J., Jinek M.;
RT "Structural basis for guide RNA processing and seed-dependent DNA targeting
RT by CRISPR-Cas12a.";
RL Mol. Cell 66:221-233(2017).
RN [9] {ECO:0007744|PDB:5MGA}
RP X-RAY CRYSTALLOGRAPHY (3.00 ANGSTROMS) IN COMPLEX WITH GUIDE RNA AND
RP PRODUCT DNA, FUNCTION AS AN ENDONUCLEASE, COFACTOR, DOMAIN, MUTAGENESIS OF
RP GLY-608; PRO-663; ASN-666; LYS-667; LYS-671; LYS-677; ARG-692; HIS-694;
RP GLU-1006; 1065-LYS-LYS-1066 AND ARG-1218, DNA-BINDING, AND RNA-BINDING.
RC STRAIN=U112;
RX PubMed=28562584; DOI=10.1038/nature22398;
RA Stella S., Alcon P., Montoya G.;
RT "Structure of the Cpf1 endonuclease R-loop complex after target DNA
RT cleavage.";
RL Nature 546:559-563(2017).
RN [10]
RP ERRATUM OF PUBMED:28562584.
RX PubMed=28678773; DOI=10.1038/nature23300;
RA Stella S., Alcon P., Montoya G.;
RT "Structure of the Cpf1 endonuclease R-loop complex after target DNA
RT cleavage.";
RL Nature 547:476-476(2017).
CC -!- FUNCTION: CRISPR (clustered regularly interspaced short palindromic
CC repeat), is an adaptive immune system that provides protection against
CC mobile genetic elements (viruses, transposable elements and conjugative
CC plasmids). CRISPR clusters contain sequences complementary to
CC antecedent mobile elements and target invading nucleic acids. CRISPR
CC clusters are transcribed and processed into CRISPR RNA (crRNA). Has
CC endonuclease activity on pre-crRNA and dsDNA, using different active
CC sites. A single-RNA guided endonuclease that is also capable of guiding
CC crRNA processing; correct processing of pre-crRNA requires only this
CC protein and the CRISPR locus (PubMed:26422227, PubMed:27096362). pre-
CC crRNA processing proceeds by an intramolecular nucleophilic attack on
CC the scissile phosphate by the 2'-OH of the upstream ribonucleotide, the
CC divalent cation (which is bound by the crRNA) is probably required for
CC ordering the crRNA pseudoknot and/or increasing RNA binding
CC (PubMed:28431230). RNA mutagenesis studies show pre-crRNA cleavage is
CC highly sequence- and structure-specific (PubMed:27096362). Forms a
CC complex with crRNA and complementary dsDNA, where the crRNA displaces
CC the non-target DNA strand and directs endonucleolytic cleavage of both
CC strands of the DNA (PubMed:26422227, PubMed:27096362, PubMed:28431230).
CC Cleavage results in staggered 5-base 5' overhangs 14-18 and 21-23 bases
CC downstream of the PAM (protospacer adjacent motif) on the non-target
CC and target strands respectively (PubMed:26422227, PubMed:28431230,
CC PubMed:28562584). Both target and non-target strand DNA are probably
CC independently cleaved in the same active site (PubMed:28431230,
CC PubMed:28562584). When this protein is expressed in E.coli it prevents
CC plasmids homologous to the first CRISPR spacer from transforming,
CC formally showing it is responsible for plasmid immunity
CC (PubMed:26422227). {ECO:0000269|PubMed:26422227,
CC ECO:0000269|PubMed:27096362, ECO:0000269|PubMed:28431230,
CC ECO:0000269|PubMed:28562584}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=Endonucleolytic cleavage to 5'-phosphodinucleotide and 5'-
CC phosphooligonucleotide end-products.; EC=3.1.21.1;
CC Evidence={ECO:0000269|PubMed:28431230};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=RNA = a 5'-hydroxy-ribonucleotide + n nucleoside-2',3'-
CC cyclophosphates.; EC=4.6.1.22;
CC Evidence={ECO:0000269|PubMed:28431230};
CC -!- COFACTOR:
CC Name=Ca(2+); Xref=ChEBI:CHEBI:29108;
CC Evidence={ECO:0000269|PubMed:26593719};
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Evidence={ECO:0000269|PubMed:26422227, ECO:0000269|PubMed:26593719,
CC ECO:0000269|PubMed:27096362, ECO:0000269|PubMed:28562584};
CC Note=Cleavage of dsDNA requires Mg(2+) (PubMed:26422227,
CC PubMed:28562584). Another report shows DNA cleavage occurs equally well
CC in the presence of Ca(2+) or Mg(2+) (PubMed:27096362). Processing of
CC pre-crRNA requires a divalent cation, preferably Mg(2+) which is bound
CC by the crRNA (PubMed:26593719, PubMed:28431230).
CC {ECO:0000269|PubMed:26422227, ECO:0000269|PubMed:26593719,
CC ECO:0000269|PubMed:27096362, ECO:0000269|PubMed:28431230,
CC ECO:0000269|PubMed:28562584};
CC -!- SUBUNIT: Might be a homodimer (PubMed:26422227). Might be a monomer
CC (PubMed:27096362, PubMed:28431230). {ECO:0000269|PubMed:26422227,
CC ECO:0000269|PubMed:27096362, ECO:0000269|PubMed:28431230}.
CC -!- DOMAIN: Has bilobed structure, with the REC lobe (residues 25-591)
CC connected to the NUC lobe (662-1300) by a discontinuous wedge domain
CC (PubMed:28431230, PubMed:28562584). The REC lobe binds the (pre-)crRNA
CC and the crRNA-target DNA heteroduplex (PubMed:28431230,
CC PubMed:28562584). The heteroduplex as well as part of the DNA
CC downstream of the heteroduplex is protected in the central cavity
CC formed by the NUC and REC lobes, which also positions target and non-
CC target DNA for cleavage after domain rearrangement (PubMed:28431230,
CC PubMed:28562584). The LKL region (residues 662 to 679) inserts into
CC target dsDNA initiating its disruption to allow crRNA hybridization, is
CC also involved in determining the non-target strand cleavage site
CC (PubMed:28562584). A 'septum' formed by residues 197-204 and 1061-1070
CC separates the 2 DNA strands, preventing their reannealing, this region
CC also influences the non-target cleavage site (PubMed:28562584).
CC {ECO:0000269|PubMed:28431230, ECO:0000269|PubMed:28562584}.
CC -!- BIOTECHNOLOGY: This class of CRISPR enzymes recognize a 5' T-rich
CC protospacer adjacent motif (PAM, TTN for this specific enzyme), unlike
CC Cas9 enzymes which recognize 3' G-rich PAMs, thus this enzyme increases
CC the possibilites for genome editing (PubMed:26422227). The simplicity
CC of the Cas12a-crRNA directed DNA endonuclease activity has been used to
CC target and modify DNA sequences in rice and tobacco (PubMed:27905529).
CC {ECO:0000269|PubMed:27272384, ECO:0000269|PubMed:27905529,
CC ECO:0000305|PubMed:26422227}.
CC -!- MISCELLANEOUS: Part of a type V-A CRISPR-Cas system.
CC {ECO:0000305|PubMed:26593719, ECO:0000305|PubMed:28111461}.
CC -!- SIMILARITY: Belongs to the CRISPR-associated endonuclease Cas12a
CC family. {ECO:0000305|PubMed:28111461}.
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DR EMBL; CP000439; ABK90267.1; -; Genomic_DNA.
DR RefSeq; WP_003040289.1; NZ_CP009633.1.
DR PDB; 5MGA; X-ray; 3.00 A; A=1-1300.
DR PDB; 5NFV; X-ray; 2.50 A; A=2-1300.
DR PDB; 5NG6; X-ray; 3.34 A; A/C/E/G=2-1300.
DR PDB; 6GTC; EM; 3.91 A; A=1-1300.
DR PDB; 6GTD; EM; 4.24 A; A=1-1300.
DR PDB; 6GTE; EM; 4.07 A; A=1-1300.
DR PDB; 6GTF; EM; 3.63 A; A=1-1300.
DR PDB; 6GTG; EM; 3.27 A; A=1-1300.
DR PDB; 6I1K; X-ray; 2.65 A; A=2-1300.
DR PDB; 6I1L; X-ray; 2.98 A; A/D=2-1300.
DR PDBsum; 5MGA; -.
DR PDBsum; 5NFV; -.
DR PDBsum; 5NG6; -.
DR PDBsum; 6GTC; -.
DR PDBsum; 6GTD; -.
DR PDBsum; 6GTE; -.
DR PDBsum; 6GTF; -.
DR PDBsum; 6GTG; -.
DR PDBsum; 6I1K; -.
DR PDBsum; 6I1L; -.
DR AlphaFoldDB; A0Q7Q2; -.
DR SMR; A0Q7Q2; -.
DR PRIDE; A0Q7Q2; -.
DR EnsemblBacteria; ABK90267; ABK90267; FTN_1397.
DR KEGG; ftn:FTN_1397; -.
DR OMA; QIYNKDF; -.
DR OrthoDB; 105944at2; -.
DR BioCyc; FTUL401614:G1G75-1444-MON; -.
DR Proteomes; UP000000762; Chromosome.
DR GO; GO:0033898; F:Bacillus subtilis ribonuclease activity; IEA:UniProtKB-EC.
DR GO; GO:0004530; F:deoxyribonuclease I activity; IEA:UniProtKB-EC.
DR GO; GO:0003677; F:DNA binding; IEA:UniProtKB-KW.
DR GO; GO:0016829; F:lyase activity; IEA:UniProtKB-KW.
DR GO; GO:0003723; F:RNA binding; IEA:UniProtKB-KW.
DR GO; GO:0051607; P:defense response to virus; IEA:UniProtKB-KW.
DR InterPro; IPR027620; Cas12a.
DR InterPro; IPR040882; Cas12a_NUC.
DR InterPro; IPR040787; Cas12a_REC1.
DR InterPro; IPR040852; RuvC_1.
DR Pfam; PF18510; NUC; 1.
DR Pfam; PF18501; REC1; 1.
DR Pfam; PF18516; RuvC_1; 1.
DR TIGRFAMs; TIGR04330; cas_Cpf1; 1.
PE 1: Evidence at protein level;
KW 3D-structure; Antiviral defense; Calcium; DNA-binding; Endonuclease;
KW Hydrolase; Lyase; Magnesium; Nuclease; RNA-binding.
FT CHAIN 1..1300
FT /note="CRISPR-associated endonuclease Cas12a"
FT /id="PRO_0000434902"
FT REGION 1..24
FT /note="Wedge region 1"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 25..339
FT /note="Recognition domain 1"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 47..51
FT /note="Binds crRNA alone and in crRNA-target DNA
FT heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT REGION 182..186
FT /note="Binds crRNA alone and in crRNA-target DNA
FT heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT REGION 301..305
FT /note="Binds DNA in crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230"
FT REGION 326..329
FT /note="Binds crRNA in crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT REGION 340..591
FT /note="Recognition domain 2"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 538..541
FT /note="Binds crRNA in crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230"
FT REGION 591..595
FT /note="Binds crRNA"
FT /evidence="ECO:0000269|PubMed:28431230"
FT REGION 592..662
FT /note="Wedge region 2"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 662..679
FT /note="LKL, important for PAM recognition and DNA
FT unwinding"
FT /evidence="ECO:0000305|PubMed:28562584"
FT REGION 663..762
FT /note="PAM-interacting domain (PI)"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 671..677
FT /note="Binds DNA protospacer adjacent motif (PAM) on target
FT DNA"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT REGION 692..704
FT /note="Binds single-strand non-target DNA"
FT /evidence="ECO:0000269|PubMed:28431230"
FT REGION 763..892
FT /note="Wedge region 3"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 791..794
FT /note="Binds crRNA"
FT /evidence="ECO:0000269|PubMed:28431230"
FT REGION 803..804
FT /note="Binds crRNA"
FT /evidence="ECO:0000269|PubMed:28431230"
FT REGION 851..853
FT /note="Binds crRNA"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT REGION 865..873
FT /note="Binds crRNA"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT REGION 893..953
FT /note="RuvC-I"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 954..971
FT /note="Bridge helix"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 972..1078
FT /note="RuvC-II"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 1079..1254
FT /note="Nuclease domain"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT REGION 1255..1300
FT /note="RuvC-III"
FT /evidence="ECO:0000303|PubMed:28431230,
FT ECO:0000303|PubMed:28562584"
FT ACT_SITE 843
FT /note="For pre-crRNA processing"
FT /evidence="ECO:0000305|PubMed:28431230"
FT ACT_SITE 852
FT /note="For pre-crRNA processing"
FT /evidence="ECO:0000305|PubMed:28431230"
FT ACT_SITE 869
FT /note="For pre-crRNA processing"
FT /evidence="ECO:0000305|PubMed:28431230"
FT ACT_SITE 917
FT /note="For DNase activity of RuvC domain"
FT /evidence="ECO:0000305|PubMed:26422227"
FT ACT_SITE 1006
FT /note="For DNase activity of RuvC domain"
FT /evidence="ECO:0000305|PubMed:26422227"
FT ACT_SITE 1255
FT /note="For DNase activity of RuvC domain"
FT /evidence="ECO:0000305|PubMed:26422227"
FT SITE 16
FT /note="Binds crRNA alone and in crRNA-target DNA
FT heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT SITE 131
FT /note="Binds target strand DNA"
FT /evidence="ECO:0000269|PubMed:28431230"
FT SITE 295
FT /note="Binds crRNA in crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230"
FT SITE 320
FT /note="Binds DNA in crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230"
FT SITE 334
FT /note="Binds DNA in crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230"
FT SITE 410
FT /note="Caps the crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000305|PubMed:28431230"
FT SITE 589
FT /note="Binds DNA in crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230"
FT SITE 613
FT /note="Binds DNA protospacer adjacent motif (PAM)"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT SITE 667
FT /note="Binds Target strand DNA"
FT /evidence="ECO:0000269|PubMed:28431230"
FT SITE 671
FT /note="Binds PAM"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT SITE 677
FT /note="Binds Target strand DNA"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT SITE 823
FT /note="Binds Target strand DNA"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT SITE 826
FT /note="Binds Target strand DNA; via amide nitrogen"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT SITE 833
FT /note="Binds crRNA"
FT /evidence="ECO:0000269|PubMed:28431230"
FT SITE 852
FT /note="Stabilizes transition state for pre-crRNA
FT processing"
FT /evidence="ECO:0000305|PubMed:28431230"
FT SITE 1026
FT /note="Binds DNA in crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT SITE 1063
FT /note="Binds DNA in crRNA-target DNA heteroduplex"
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT MUTAGEN 608
FT /note="G->A,E: 15% DNA cleavage."
FT /evidence="ECO:0000269|PubMed:28562584"
FT MUTAGEN 663
FT /note="P->A: 25% DNA cleavage, altered non-target strand
FT cleavage products."
FT /evidence="ECO:0000269|PubMed:28562584"
FT MUTAGEN 666
FT /note="N->A: 80% DNA cleavage, altered non-target strand
FT cleavage products."
FT /evidence="ECO:0000269|PubMed:28562584"
FT MUTAGEN 667
FT /note="K->A: 30% DNA cleavage."
FT /evidence="ECO:0000269|PubMed:28562584"
FT MUTAGEN 671
FT /note="K->A: 15% DNA cleavage."
FT /evidence="ECO:0000269|PubMed:28562584"
FT MUTAGEN 677
FT /note="K->A: 35% DNA cleavage, altered non-target strand
FT cleavage products."
FT /evidence="ECO:0000269|PubMed:28562584"
FT MUTAGEN 692
FT /note="R->A: Slight decrease in target DNA cleavage, 30%
FT DNA cleavage, altered non-target strand cleavage products."
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT MUTAGEN 694
FT /note="H->A: Wild-type DNA cleavage, altered non-target
FT strand cleavage products."
FT /evidence="ECO:0000269|PubMed:28562584"
FT MUTAGEN 698..702
FT /note="TKNGS->AGGGG: Loss of target DNA cleavage."
FT /evidence="ECO:0000269|PubMed:28431230"
FT MUTAGEN 704
FT /note="Q->A: Significant decrease in target DNA cleavage."
FT /evidence="ECO:0000269|PubMed:28431230"
FT MUTAGEN 843
FT /note="H->A: Decreased pre-crRNA processing in vitro, binds
FT RNA, no change in DNA cleavage."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 852
FT /note="K->A: Decreased pre-crRNA processing in vitro, binds
FT RNA, no change in DNA cleavage."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 869
FT /note="K->A: Decreased pre-crRNA processing in vitro, binds
FT RNA, no change in DNA cleavage."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 873
FT /note="F->A: Decreased pre-crRNA processing in vitro, no
FT pre-crRNA processing in E.coli, binds RNA, no change in DNA
FT cleavage."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 917
FT /note="D->A: Loss of target and non-target strand DNA
FT cleavage, no change in DNA-binding or pre-crRNA
FT processing."
FT /evidence="ECO:0000269|PubMed:26422227,
FT ECO:0000269|PubMed:27096362, ECO:0000269|PubMed:28431230"
FT MUTAGEN 920
FT /note="E->A: No longer cleaves DNA in presence of Ca(2+)."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 922
FT /note="H->A: Decreased cleavage of target strand in
FT presence of Ca(2+), wild-type cleavage of DNA in presence
FT of Mg(2+)."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 925
FT /note="Y->A: Decreased cleavage of target strand in
FT presence of Ca(2+), wild-type cleavage of DNA in presence
FT of Mg(2+)."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 1006
FT /note="E->A: Loss of target and non-target strand DNA
FT cleavage, no change in DNA-binding or pre-crRNA
FT processing."
FT /evidence="ECO:0000269|PubMed:26422227,
FT ECO:0000269|PubMed:27096362, ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT MUTAGEN 1006
FT /note="E->Q: Complete loss of DNA cleavage, still binds
FT crRNA; when associated with A-1218."
FT /evidence="ECO:0000269|PubMed:28431230"
FT MUTAGEN 1024
FT /note="Y->A: No longer cleaves DNA in presence of Ca(2+)."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 1028
FT /note="E->A: No longer cleaves DNA in presence of Ca(2+),
FT reduced cleavage of non-target strand in presence of
FT Mg(2+)."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 1065..1066
FT /note="KK->AA: 67% DNA cleavage, altered non-target strand
FT cleavage products."
FT /evidence="ECO:0000269|PubMed:28562584"
FT MUTAGEN 1218
FT /note="R->A: Cleaves both target and non-target strand DNA.
FT Complete loss of DNA cleavage, still binds crRNA; when
FT associated with Q-1006."
FT /evidence="ECO:0000269|PubMed:28431230,
FT ECO:0000269|PubMed:28562584"
FT MUTAGEN 1227
FT /note="D->A: No longer cleaves DNA in presence of Ca(2+)."
FT /evidence="ECO:0000269|PubMed:27096362"
FT MUTAGEN 1255
FT /note="D->A: Significant reduction to loss of target and
FT non-target strand DNA cleavage, no change in DNA-binding or
FT pre-crRNA processing."
FT /evidence="ECO:0000269|PubMed:26422227,
FT ECO:0000269|PubMed:27096362, ECO:0000269|PubMed:28431230"
FT MUTAGEN 1255
FT /note="D->N: Significant reduction of target and non-target
FT strand DNA cleavage."
FT /evidence="ECO:0000269|PubMed:28431230"
FT TURN 3..6
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 13..23
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 27..34
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 36..68
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 73..86
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 92..114
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 117..120
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 122..124
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 125..127
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 132..134
FT /evidence="ECO:0007829|PDB:6I1L"
FT HELIX 137..147
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 153..155
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 157..159
FT /evidence="ECO:0007829|PDB:6I1K"
FT HELIX 162..171
FT /evidence="ECO:0007829|PDB:5NFV"
FT TURN 172..174
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 176..179
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 180..190
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 192..194
FT /evidence="ECO:0007829|PDB:6I1K"
FT STRAND 196..198
FT /evidence="ECO:0007829|PDB:5NG6"
FT HELIX 199..204
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 207..224
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 226..228
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 231..237
FT /evidence="ECO:0007829|PDB:5NFV"
FT TURN 238..242
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 243..245
FT /evidence="ECO:0007829|PDB:5NFV"
FT TURN 248..251
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 252..257
FT /evidence="ECO:0007829|PDB:6I1L"
FT HELIX 260..263
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 267..271
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 272..274
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 275..286
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 291..293
FT /evidence="ECO:0007829|PDB:5MGA"
FT STRAND 294..297
FT /evidence="ECO:0007829|PDB:5NG6"
FT HELIX 300..311
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 314..319
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 345..361
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 365..367
FT /evidence="ECO:0007829|PDB:6I1K"
FT HELIX 370..382
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 388..390
FT /evidence="ECO:0007829|PDB:5NG6"
FT STRAND 392..394
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 397..406
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 412..422
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 424..426
FT /evidence="ECO:0007829|PDB:6GTG"
FT STRAND 427..432
FT /evidence="ECO:0007829|PDB:6I1L"
FT HELIX 435..445
FT /evidence="ECO:0007829|PDB:6I1K"
FT STRAND 449..452
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 453..465
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 469..471
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 475..484
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 487..506
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 508..510
FT /evidence="ECO:0007829|PDB:6I1L"
FT HELIX 513..515
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 517..519
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 520..541
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 559..572
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 575..586
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 595..597
FT /evidence="ECO:0007829|PDB:5NFV"
FT TURN 603..606
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 611..613
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 614..617
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 619..624
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 627..633
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 635..637
FT /evidence="ECO:0007829|PDB:6I1K"
FT TURN 638..641
FT /evidence="ECO:0007829|PDB:6I1K"
FT HELIX 643..648
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 650..661
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 665..673
FT /evidence="ECO:0007829|PDB:5NFV"
FT TURN 676..678
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 679..682
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 686..694
FT /evidence="ECO:0007829|PDB:5NFV"
FT TURN 695..697
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 698..700
FT /evidence="ECO:0007829|PDB:6I1L"
FT HELIX 704..706
FT /evidence="ECO:0007829|PDB:6I1L"
FT HELIX 714..730
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 731..733
FT /evidence="ECO:0007829|PDB:6I1K"
FT HELIX 734..737
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 744..746
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 750..760
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 761..769
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 771..779
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 782..789
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 791..793
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 803..812
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 814..818
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 821..824
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 829..833
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 845..847
FT /evidence="ECO:0007829|PDB:6GTG"
FT STRAND 852..856
FT /evidence="ECO:0007829|PDB:5MGA"
FT STRAND 857..861
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 871..874
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 877..887
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 888..890
FT /evidence="ECO:0007829|PDB:6GTG"
FT HELIX 896..906
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 908..910
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 912..917
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 919..921
FT /evidence="ECO:0007829|PDB:6GTG"
FT STRAND 923..929
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 931..933
FT /evidence="ECO:0007829|PDB:6I1K"
FT STRAND 935..944
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 947..952
FT /evidence="ECO:0007829|PDB:6I1K"
FT HELIX 953..968
FT /evidence="ECO:0007829|PDB:5NFV"
FT TURN 969..971
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 977..999
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1001..1006
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1008..1011
FT /evidence="ECO:0007829|PDB:6I1K"
FT HELIX 1016..1018
FT /evidence="ECO:0007829|PDB:6GTG"
FT HELIX 1019..1036
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1041..1043
FT /evidence="ECO:0007829|PDB:5MGA"
FT STRAND 1045..1047
FT /evidence="ECO:0007829|PDB:6I1K"
FT STRAND 1050..1053
FT /evidence="ECO:0007829|PDB:6I1K"
FT HELIX 1065..1067
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1069..1071
FT /evidence="ECO:0007829|PDB:6I1L"
FT STRAND 1074..1077
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1083..1085
FT /evidence="ECO:0007829|PDB:5NFV"
FT TURN 1087..1089
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 1102..1110
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1112..1118
FT /evidence="ECO:0007829|PDB:5NFV"
FT TURN 1119..1122
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1123..1129
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 1130..1132
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1133..1135
FT /evidence="ECO:0007829|PDB:6GTG"
FT STRAND 1136..1138
FT /evidence="ECO:0007829|PDB:6I1L"
FT STRAND 1141..1145
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1150..1153
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1157..1162
FT /evidence="ECO:0007829|PDB:6GTG"
FT STRAND 1165..1168
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 1170..1180
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 1192..1197
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 1201..1214
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1218..1220
FT /evidence="ECO:0007829|PDB:6I1L"
FT TURN 1222..1225
FT /evidence="ECO:0007829|PDB:5NG6"
FT STRAND 1228..1230
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1236..1238
FT /evidence="ECO:0007829|PDB:6I1K"
FT STRAND 1242..1245
FT /evidence="ECO:0007829|PDB:5NFV"
FT HELIX 1254..1275
FT /evidence="ECO:0007829|PDB:5NFV"
FT STRAND 1278..1280
FT /evidence="ECO:0007829|PDB:6I1K"
FT HELIX 1288..1297
FT /evidence="ECO:0007829|PDB:5NFV"
SQ SEQUENCE 1300 AA; 151915 MW; 601E903DE68C80DE CRC64;
MSIYQEFVNK YSLSKTLRFE LIPQGKTLEN IKARGLILDD EKRAKDYKKA KQIIDKYHQF
FIEEILSSVC ISEDLLQNYS DVYFKLKKSD DDNLQKDFKS AKDTIKKQIS EYIKDSEKFK
NLFNQNLIDA KKGQESDLIL WLKQSKDNGI ELFKANSDIT DIDEALEIIK SFKGWTTYFK
GFHENRKNVY SSNDIPTSII YRIVDDNLPK FLENKAKYES LKDKAPEAIN YEQIKKDLAE
ELTFDIDYKT SEVNQRVFSL DEVFEIANFN NYLNQSGITK FNTIIGGKFV NGENTKRKGI
NEYINLYSQQ INDKTLKKYK MSVLFKQILS DTESKSFVID KLEDDSDVVT TMQSFYEQIA
AFKTVEEKSI KETLSLLFDD LKAQKLDLSK IYFKNDKSLT DLSQQVFDDY SVIGTAVLEY
ITQQIAPKNL DNPSKKEQEL IAKKTEKAKY LSLETIKLAL EEFNKHRDID KQCRFEEILA
NFAAIPMIFD EIAQNKDNLA QISIKYQNQG KKDLLQASAE DDVKAIKDLL DQTNNLLHKL
KIFHISQSED KANILDKDEH FYLVFEECYF ELANIVPLYN KIRNYITQKP YSDEKFKLNF
ENSTLANGWD KNKEPDNTAI LFIKDDKYYL GVMNKKNNKI FDDKAIKENK GEGYKKIVYK
LLPGANKMLP KVFFSAKSIK FYNPSEDILR IRNHSTHTKN GSPQKGYEKF EFNIEDCRKF
IDFYKQSISK HPEWKDFGFR FSDTQRYNSI DEFYREVENQ GYKLTFENIS ESYIDSVVNQ
GKLYLFQIYN KDFSAYSKGR PNLHTLYWKA LFDERNLQDV VYKLNGEAEL FYRKQSIPKK
ITHPAKEAIA NKNKDNPKKE SVFEYDLIKD KRFTEDKFFF HCPITINFKS SGANKFNDEI
NLLLKEKAND VHILSIDRGE RHLAYYTLVD GKGNIIKQDT FNIIGNDRMK TNYHDKLAAI
EKDRDSARKD WKKINNIKEM KEGYLSQVVH EIAKLVIEYN AIVVFEDLNF GFKRGRFKVE
KQVYQKLEKM LIEKLNYLVF KDNEFDKTGG VLRAYQLTAP FETFKKMGKQ TGIIYYVPAG
FTSKICPVTG FVNQLYPKYE SVSKSQEFFS KFDKICYNLD KGYFEFSFDY KNFGDKAAKG
KWTIASFGSR LINFRNSDKN HNWDTREVYP TKELEKLLKD YSIEYGHGEC IKAAICGESD
KKFFAKLTSV LNTILQMRNS KTGTELDYLI SPVADVNGNF FDSRQAPKNM PQDADANGAY
HIGLKGLMLL GRIKNNQEGK KLNLVIKNEE YFEFVQNRNN
