P45956 · CAS2_ECOLI
- ProteinCRISPR-associated endoribonuclease Cas2
- GeneygbF
- StatusUniProtKB reviewed (Swiss-Prot)
- Organism
- Amino acids
- Protein existenceEvidence at protein level
- Annotation score5/5
Function
function
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) (PubMed:21255106, PubMed:24793649, PubMed:24920831).
CRISPR clusters contain sequences complementary to antecedent mobile elements and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA). The Cas1-Cas2 complex is involved in CRISPR adaptation, the first stage of CRISPR immunity, being required for the addition/removal of CRISPR spacers at the leader end of the CRISPR locus (PubMed:24793649, PubMed:24920831, PubMed:25707795).
The Cas1-Cas2 complex introduces staggered nicks into both strands of the CRISPR array near the leader repeat and joins the 5'-ends of the repeat strands with the 3'-ends of the new spacer sequence (PubMed:24920831).
Spacer DNA integration requires supercoiled target DNA and 3'-OH ends on the inserted (spacer) DNA and probably initiates with a nucleophilic attack of the C 3'-OH end of the protospacer on the minus strand of the first repeat sequence (PubMed:25707795).
Expression of Cas1-Cas2 in a strain lacking both genes permits spacer acquisition (PubMed:24793649, PubMed:24920831).
Cas2 not seen to bind DNA alone; the Cas1-Cas2 complex preferentially binds CRISPR-locus DNA (PubMed:24793649).
Highest binding is seen to a dual forked DNA complex with 3'-overhangs and a protospacer-adjacent motif-complement specifically positioned (PubMed:26478180).
The protospacer DNA lies across a flat surface extending from 1 Cas1 dimer, across the Cas2 dimer and contacting the other Cas1 dimer; the 23 bp-long ds section of the DNA is bracketed by 1 Tyr-22 from each of the Cas1 dimers (PubMed:26478180, PubMed:26503043).
Cas1 cuts within the 3'-overhang, to generate a 33-nucleotide DNA that is probably incorporated into the CRISPR leader by a cut-and-paste mechanism (PubMed:26478180).
This subunit's probable nuclease activity is not required for spacer acquisition (PubMed:24793649).
CRISPR clusters contain sequences complementary to antecedent mobile elements and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA). The Cas1-Cas2 complex is involved in CRISPR adaptation, the first stage of CRISPR immunity, being required for the addition/removal of CRISPR spacers at the leader end of the CRISPR locus (PubMed:24793649, PubMed:24920831, PubMed:25707795).
The Cas1-Cas2 complex introduces staggered nicks into both strands of the CRISPR array near the leader repeat and joins the 5'-ends of the repeat strands with the 3'-ends of the new spacer sequence (PubMed:24920831).
Spacer DNA integration requires supercoiled target DNA and 3'-OH ends on the inserted (spacer) DNA and probably initiates with a nucleophilic attack of the C 3'-OH end of the protospacer on the minus strand of the first repeat sequence (PubMed:25707795).
Expression of Cas1-Cas2 in a strain lacking both genes permits spacer acquisition (PubMed:24793649, PubMed:24920831).
Cas2 not seen to bind DNA alone; the Cas1-Cas2 complex preferentially binds CRISPR-locus DNA (PubMed:24793649).
Highest binding is seen to a dual forked DNA complex with 3'-overhangs and a protospacer-adjacent motif-complement specifically positioned (PubMed:26478180).
The protospacer DNA lies across a flat surface extending from 1 Cas1 dimer, across the Cas2 dimer and contacting the other Cas1 dimer; the 23 bp-long ds section of the DNA is bracketed by 1 Tyr-22 from each of the Cas1 dimers (PubMed:26478180, PubMed:26503043).
Cas1 cuts within the 3'-overhang, to generate a 33-nucleotide DNA that is probably incorporated into the CRISPR leader by a cut-and-paste mechanism (PubMed:26478180).
This subunit's probable nuclease activity is not required for spacer acquisition (PubMed:24793649).
GO annotations
Aspect | Term | |
---|---|---|
Molecular Function | DNA binding | |
Molecular Function | endonuclease activity | |
Biological Process | CRISPR-cas system | |
Biological Process | defense response to virus | |
Biological Process | maintenance of CRISPR repeat elements |
Keywords
- Molecular function
- Biological process
Enzyme and pathway databases
Names & Taxonomy
Protein names
- Recommended nameCRISPR-associated endoribonuclease Cas2
- EC number
Gene names
Organism names
- Organism
- Strains
- Taxonomic lineageBacteria > Pseudomonadota > Gammaproteobacteria > Enterobacterales > Enterobacteriaceae > Escherichia
Accessions
- Primary accessionP45956
- Secondary accessions
Proteomes
Phenotypes & Variants
Disruption phenotype
Loss of plasmid silencing (PubMed:21255106).
Features
Showing features for mutagenesis.
Type | ID | Position(s) | Description | |||
---|---|---|---|---|---|---|
Mutagenesis | 9 | No effect on spacer acquisition, Cas1-Cas2 complex formation or CRISPRDNA-binding by complex. | ||||
Sequence: E → A or R | ||||||
Mutagenesis | 10 | No effect on spacer acquisition. | ||||
Sequence: N → A | ||||||
Mutagenesis | 14 | Slight decrease in spacer acquisition. | ||||
Sequence: R → A | ||||||
Mutagenesis | 14-16 | No in vivspacer acquisition, significantly decreased protospacer binding. | ||||
Sequence: RLR → ALA | ||||||
Mutagenesis | 16 | Slight decrease in spacer acquisition. | ||||
Sequence: R → A | ||||||
Mutagenesis | 16 | Dramatically decreased spacer acquisition in vivo. | ||||
Sequence: R → E | ||||||
Mutagenesis | 18 | Very little spacer acquisition. | ||||
Sequence: R → A | ||||||
Mutagenesis | 27 | Slight decrease in spacer acquisition. | ||||
Sequence: R → A | ||||||
Mutagenesis | 38-40 | Very little in vivo spacer acquisition. | ||||
Sequence: KIR → AIA | ||||||
Mutagenesis | 65 | No effect on spacer acquisition. | ||||
Sequence: E → A | ||||||
Mutagenesis | 65 | Slight decrease in spacer acquisition, Cas1-Cas2 complex formation or CRISPRDNA-binding by complex. Loss of spacer acquisition; when associated with R-84. | ||||
Sequence: E → R | ||||||
Mutagenesis | 77 | No change in spacer acquisition in vivo. | ||||
Sequence: R → E | ||||||
Mutagenesis | 77-78 | No spacer acquisition, significantly decreased protospacer binding. | ||||
Sequence: RR → AA | ||||||
Mutagenesis | 78 | Dramatically decreased spacer acquisition in vivo. | ||||
Sequence: R → E | ||||||
Mutagenesis | 79-94 | Loss of spacer acquisition, no Cas1-Cas2 complex formation, loss of CRISPRDNA-binding by complex (beta6-beta7 deletion). | ||||
Sequence: Missing | ||||||
Mutagenesis | 84 | No effect on spacer acquisition. | ||||
Sequence: D → A | ||||||
Mutagenesis | 84 | Slight decrease in spacer acquisition. Loss of spacer acquisition; when associated with R-65. | ||||
Sequence: D → R |
PTM/Processing
Features
Showing features for chain.
Type | ID | Position(s) | Description | |||
---|---|---|---|---|---|---|
Chain | PRO_0000169312 | 1-94 | CRISPR-associated endoribonuclease Cas2 | |||
Sequence: MSMLVVVTENVPPRLRGRLAIWLLEVRAGVYVGDVSAKIREMIWEQIAGLAEEGNVVMAWATNTETGFEFQTFGLNRRTPVDLDGLRLVSFLPV |
Proteomic databases
Interaction
Subunit
Homodimer (Ref.10). Part of the Cas1-Cas2 complex (PubMed:24793649, PubMed:24920831, PubMed:25707795, PubMed:26478180, PubMed:26503043, Ref.12). Forms a hexamer with 2 Cas1 dimers sandwiching a Cas2 dimer (PubMed:24793649).
The DNA lies across a flat surface extending from 1 Cas1 dimer, across the Cas2 dimer and contacting the other Cas1 dimer. Only 1 Cas1 protein from each dimer is catalytic, the other interacts with the Cas2 dimer and possibly target DNA (PubMed:26478180, PubMed:26503043).
The DNA lies across a flat surface extending from 1 Cas1 dimer, across the Cas2 dimer and contacting the other Cas1 dimer. Only 1 Cas1 protein from each dimer is catalytic, the other interacts with the Cas2 dimer and possibly target DNA (PubMed:26478180, PubMed:26503043).
Binary interactions
Type | Entry 1 | Entry 2 | Number of experiments | Intact | |
---|---|---|---|---|---|
BINARY | P45956 | ygbT Q46896 | 8 | EBI-9150552, EBI-1130209 |
Protein-protein interaction databases
Structure
Family & Domains
Domain
Substrate DNA-binding induces large structural changes that generate a surface for DNA-binding across the Cas2 dimer and formation of an optimal catalytic site (PubMed:26478180).
Sequence similarities
Phylogenomic databases
Family and domain databases
Sequence
- Sequence statusComplete
- Length94
- Mass (Da)10,518
- Last updated2000-05-30 v2
- ChecksumD1C159D924B477B4
Keywords
- Technical term
Sequence databases
Nucleotide Sequence | Protein Sequence | Molecule Type | Status | |
---|---|---|---|---|
M27059 EMBL· GenBank· DDBJ | - | Unassigned DNA | No translation available. | |
U29579 EMBL· GenBank· DDBJ | AAA69264.1 EMBL· GenBank· DDBJ | Genomic DNA | ||
U00096 EMBL· GenBank· DDBJ | AAC75796.2 EMBL· GenBank· DDBJ | Genomic DNA | ||
AP009048 EMBL· GenBank· DDBJ | BAE76831.1 EMBL· GenBank· DDBJ | Genomic DNA |