Q46896 · CAS1_ECOLI
- ProteinCRISPR-associated endonuclease Cas1
- GeneygbT
- StatusUniProtKB reviewed (Swiss-Prot)
- Organism
- Amino acids305 (go to sequence)
- 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).
Non-specifically binds DNA; 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).
Cas1 alone endonucleolytically cleaves linear ssRNA, ssDNA and short (34 base) dsDNA as well as branched DNA substrates such as Holliday junctions, replication forks and 5'-flap DNA substrates (PubMed:21219465).
In vitro catalyzes a concerted transesterification reaction on branched DNA, as would be expected during integration of protospacers into the CRISPR leader sequence; Cas2 is not required in vitro. This reaction requires a 3'-OH group at the branch point (PubMed:26284603).
Genetic interactions suggest Cas1 interacts with components of the RecBC and RuvB DNA repair systems (PubMed:21219465).
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).
Non-specifically binds DNA; 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).
Cas1 alone endonucleolytically cleaves linear ssRNA, ssDNA and short (34 base) dsDNA as well as branched DNA substrates such as Holliday junctions, replication forks and 5'-flap DNA substrates (PubMed:21219465).
In vitro catalyzes a concerted transesterification reaction on branched DNA, as would be expected during integration of protospacers into the CRISPR leader sequence; Cas2 is not required in vitro. This reaction requires a 3'-OH group at the branch point (PubMed:26284603).
Genetic interactions suggest Cas1 interacts with components of the RecBC and RuvB DNA repair systems (PubMed:21219465).
Cofactor
Activity regulation
Nuclease activity partially inhibited by CasE (PubMed:21219465).
Features
Showing features for binding site.
GO annotations
Aspect | Term | |
---|---|---|
Cellular Component | cytoplasm | |
Molecular Function | 5'-flap endonuclease activity | |
Molecular Function | crossover junction DNA endonuclease activity | |
Molecular Function | DNA binding | |
Molecular Function | identical protein binding | |
Molecular Function | metal ion binding | |
Molecular Function | protein homodimerization activity | |
Biological Process | CRISPR-cas system | |
Biological Process | defense response to virus | |
Biological Process | DNA damage response | |
Biological Process | DNA repair | |
Biological Process | maintenance of CRISPR repeat elements |
Keywords
- Molecular function
- Biological process
- Ligand
Enzyme and pathway databases
Names & Taxonomy
Protein names
- Recommended nameCRISPR-associated endonuclease Cas1
- EC number
Gene names
Organism names
- Organism
- Strains
- Taxonomic lineageBacteria > Pseudomonadota > Gammaproteobacteria > Enterobacterales > Enterobacteriaceae > Escherichia
Accessions
- Primary accessionQ46896
- Secondary accessions
Proteomes
Phenotypes & Variants
Disruption phenotype
Not essential. Increased sensitivity to MMC and UV light; double ygbT-ruvA, ruvB or ruvC disruptions have no further phenotype suggesting Cas1 functions in the same DNA repair pathway (PubMed:21219465).
Function in DNA repair also seems to require CRISPRs (PubMed:21219465).
Cells elongate after 2 hours growth in MMC; they are even longer in double ygbT-ruvA, ruvB or ruvC disruptions, suggesting Cas1 may also function in chromosome segregation (PubMed:21219465).
Loss of plasmid silencing (PubMed:21255106).
Function in DNA repair also seems to require CRISPRs (PubMed:21219465).
Cells elongate after 2 hours growth in MMC; they are even longer in double ygbT-ruvA, ruvB or ruvC disruptions, suggesting Cas1 may also function in chromosome segregation (PubMed:21219465).
Loss of plasmid silencing (PubMed:21255106).
Features
Showing features for mutagenesis.
Type | ID | Position(s) | Description | |||
---|---|---|---|---|---|---|
Mutagenesis | 22 | Slightly decreased spacer acquisition in vivo. | ||||
Sequence: Y → A | ||||||
Mutagenesis | 22 | Nearly wild-type spacer acquisition in vivo. | ||||
Sequence: Y → F | ||||||
Mutagenesis | 41 | Dramatically decreased spacer acquisition in vivo. | ||||
Sequence: R → E | ||||||
Mutagenesis | 59 | Loss of spacer acquisition in vivo, decreased protospacer binding. | ||||
Sequence: R → A | ||||||
Mutagenesis | 59 | Dramatically decreased spacer acquisition in vitro, 250-fold decreased affinity for protospacer DNA. | ||||
Sequence: R → D | ||||||
Mutagenesis | 66 | Dramatically decreased spacer acquisition in vitro, 250-fold decreased affinity for protospacer DNA. | ||||
Sequence: R → D | ||||||
Mutagenesis | 66 | Dramatically decreased spacer acquisition in vivo. | ||||
Sequence: R → E | ||||||
Mutagenesis | 84 | Decreased spacer acquisition in vivo. | ||||
Sequence: R → A | ||||||
Mutagenesis | 84 | Dramatically decreased spacer acquisition in vivo. | ||||
Sequence: R → E | ||||||
Mutagenesis | 141 | No cleavage of any substrates, no restoration of UV or mitomycin C (MMC) resistance (PubMed:21219465). Loss of spacer acquisition in vivo (PubMed:24793649). | ||||
Sequence: E → A | ||||||
Mutagenesis | 149 | No effect on in vitro protospacer integration. | ||||
Sequence: Y → A | ||||||
Mutagenesis | 165 | No effect on in vitro protospacer integration (PubMed:25707795). Alone significantly decreased protospacer acquisition in vivo (PubMed:26478180). Loss of protospacer acquisition, decreased protospacer binding; in association with A-170, significantly decreased protospacer binding; in association with A-217 (PubMed:26478180). | ||||
Sequence: Y → A | ||||||
Mutagenesis | 170 | Alone significantly decreased protospacer acquisition in vivo (PubMed:26478180). Decreased protospacer binding; in association with A-170 (PubMed:26478180). | ||||
Sequence: W → A | ||||||
Mutagenesis | 184 | No cleavage of any substrates. | ||||
Sequence: T → A | ||||||
Mutagenesis | 188 | Partial inhibition of cleavage (PubMed:21219465). No effect on in vitro protospacer integration (PubMed:25707795). Significantly decreased protospacer acquisition in vivo (PubMed:26478180). | ||||
Sequence: Y → A | ||||||
Mutagenesis | 208 | No cleavage of any substrates, no restoration of UV or MMC resistance (PubMed:21219465). Loss of spacer acquisition in vivo (PubMed:24793649, PubMed:25707795, PubMed:26478180). | ||||
Sequence: H → A | ||||||
Mutagenesis | 211 | No cleavage of any substrates. | ||||
Sequence: K → A | ||||||
Mutagenesis | 217 | No effect on in vitro protospacer integration (PubMed:25707795). Alone significantly decreased protospacer acquisition in vivo (PubMed:26478180). Significantly decreased protospacer binding; in association with A-165 (PubMed:26478180). | ||||
Sequence: Y → A | ||||||
Mutagenesis | 218 | No cleavage of any substrates, no restoration of UV or MMC resistance (PubMed:21219465). Loss of spacer acquisition in vivo (PubMed:24793649). | ||||
Sequence: D → A | ||||||
Mutagenesis | 221 | No cleavage of any substrates (PubMed:21219465). Loss of spacer acquisition in vivo (PubMed:24793649, PubMed:24920831, PubMed:25707795). No cleavage of CRISPR leader in preparation for spacer integration (PubMed:24920831). | ||||
Sequence: D → A | ||||||
Mutagenesis | 224 | No cleavage of any substrates (PubMed:21219465). Loss of spacer acquisition in vivo (PubMed:24793649, PubMed:25707795). | ||||
Sequence: K → A | ||||||
Mutagenesis | 245 | No effect on spacer acquisition. | ||||
Sequence: R → A | ||||||
Mutagenesis | 245 | Decreased spacer acquisition. | ||||
Sequence: R → D | ||||||
Mutagenesis | 245 | Dramatically decreased spacer acquisition in vivo. | ||||
Sequence: R → E | ||||||
Mutagenesis | 245-248 | Loss of spacer acquisition in vivo. | ||||
Sequence: REVR → AEVA | ||||||
Mutagenesis | 248 | Dramatically decreased spacer acquisition in vivo. | ||||
Sequence: R → E | ||||||
Mutagenesis | 252 | No effect on spacer acquisition. | ||||
Sequence: R → A | ||||||
Mutagenesis | 252 | Loss of spacer acquisition, no Cas1-Cas2 complex formation, loss of CRISPRDNA-binding by complex. Protein is stable and dimerizes. | ||||
Sequence: R → E | ||||||
Mutagenesis | 256 | No effect on spacer acquisition. | ||||
Sequence: R → A | ||||||
Mutagenesis | 256 | Loss of spacer acquisition. | ||||
Sequence: R → E | ||||||
Mutagenesis | 256-259 | Loss of spacer acquisition in vivo. | ||||
Sequence: RSSK → ASSA | ||||||
Mutagenesis | 282-305 | No effect on spacer acquisition, Cas1-Cas2 complex formation or CRISPRDNA-binding by complex. | ||||
Sequence: Missing | ||||||
Mutagenesis | 291 | No effect on spacer acquisition. | ||||
Sequence: I → G or R |
PTM/Processing
Features
Showing features for chain.
Type | ID | Position(s) | Description | |||
---|---|---|---|---|---|---|
Chain | PRO_0000169315 | 1-305 | CRISPR-associated endonuclease Cas1 | |||
Sequence: MTWLPLNPIPLKDRVSMIFLQYGQIDVIDGAFVLIDKTGIRTHIPVGSVACIMLEPGTRVSHAAVRLAAQVGTLLVWVGEAGVRVYASGQPGGARSDKLLYQAKLALDEDLRLKVVRKMFELRFGEPAPARRSVEQLRGIEGSRVRATYALLAKQYGVTWNGRRYDPKDWEKGDTINQCISAATSCLYGVTEAAILAAGYAPAIGFVHTGKPLSFVYDIADIIKFDTVVPKAFEIARRNPGEPDREVRLACRDIFRSSKTLAKLIPLIEDVLAAGEIQPPAPPEDAQPVAIPLPVSLGDAGHRSS |
Proteomic databases
Interaction
Subunit
Homodimer (PubMed:21219465).
Part of the Cas1-Cas2 complex (PubMed:24793649, PubMed:24920831, PubMed:25707795, PubMed:26478180, PubMed:26503043, Ref.11). Interacts with RecB, RecC, RuvB, CasC and CasE (PubMed:21219465).
Forms a hexamer with 2 Cas1 dimers sandwiching a Cas2 dimer (PubMed:24793649, PubMed:26478180).
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).
Part of the Cas1-Cas2 complex (PubMed:24793649, PubMed:24920831, PubMed:25707795, PubMed:26478180, PubMed:26503043, Ref.11). Interacts with RecB, RecC, RuvB, CasC and CasE (PubMed:21219465).
Forms a hexamer with 2 Cas1 dimers sandwiching a Cas2 dimer (PubMed:24793649, PubMed:26478180).
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 | Q46896 | hprS P76339 | 3 | EBI-1130209, EBI-554869 | |
BINARY | Q46896 | ygbF P45956 | 8 | EBI-1130209, EBI-9150552 | |
BINARY | Q46896 | ygbT Q46896 | 4 | EBI-1130209, EBI-1130209 |
Protein-protein interaction databases
Structure
Family & Domains
Features
Showing features for region.
Type | ID | Position(s) | Description | |||
---|---|---|---|---|---|---|
Region | 96-278 | Sufficient for cleavage of ssRNA, ssDNA and Holliday junction DNA | ||||
Sequence: SDKLLYQAKLALDEDLRLKVVRKMFELRFGEPAPARRSVEQLRGIEGSRVRATYALLAKQYGVTWNGRRYDPKDWEKGDTINQCISAATSCLYGVTEAAILAAGYAPAIGFVHTGKPLSFVYDIADIIKFDTVVPKAFEIARRNPGEPDREVRLACRDIFRSSKTLAKLIPLIEDVLAAGEIQ | ||||||
Region | 278-305 | Disordered | ||||
Sequence: QPPAPPEDAQPVAIPLPVSLGDAGHRSS |
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
Belongs to the CRISPR-associated endonuclease Cas1 family.
Phylogenomic databases
Family and domain databases
Sequence
- Sequence statusComplete
- Length305
- Mass (Da)33,194
- Last updated1996-11-01 v1
- Checksum01A31BA98453D8A5
Keywords
- Technical term
Sequence databases
Nucleotide Sequence | Protein Sequence | Molecule Type | Status | |
---|---|---|---|---|
U29579 EMBL· GenBank· DDBJ | AAA69265.1 EMBL· GenBank· DDBJ | Genomic DNA | ||
U00096 EMBL· GenBank· DDBJ | AAC75797.1 EMBL· GenBank· DDBJ | Genomic DNA | ||
AP009048 EMBL· GenBank· DDBJ | BAE76832.1 EMBL· GenBank· DDBJ | Genomic DNA |