CRISPR-associated (Cas) protein
A new CRISPR-associated (Cas) protein, termed “CasM,” is described, as well as polynucleotides encoding the same and methods of using CasM for site-specific genome engineering. CasM proteins are capable of targeting and cleaving single-stranded RNA.
1. A Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated (Cas) nucleoprotein complex, wherein the nucleoprotein complex comprises a Cas protein selected from the group consisting of SEQ ID NO:37, SEQ ID NO:39, and SEQ ID NO:44, an amino acid sequence having at least 98 percent sequence identity to SEQ ID NO:37, an amino acid sequence having at least 98 percent sequence identity to SEQ ID NO:39, and an amino acid sequence having at least 98 percent sequence identity to SEQ ID NO:44; and a cognate nucleic acid guide comprising a repeat sequence and a spacer sequence, wherein the repeat sequence and the spacer sequence do not naturally occur together, wherein the Cas protein and the cognate nucleic acid guide form the nucleoprotein complex, and wherein the nucleoprotein complex is capable of site-directed binding to a target nucleic acid sequence.
2. The Cas nucleoprotein complex of claim 1 , wherein the cognate nucleic acid guide comprises a crRNA.
3. The Cas nucleoprotein complex of claim 1 , wherein the Cas protein comprises an N- or C-terminal nuclear localization signal sequence (NLS).
4. The Cas nucleoprotein complex of claim 1 , wherein the Cas protein comprises an amino acid sequence having at least 98% sequence identity to SEQ ID NO:44.
5. The Cas nucleoprotein complex of claim 1 , wherein the cognate nucleic acid guide comprises a modified base analog.
6. A eukaryotic cell comprising the Cas nucleoprotein complex of claim 1 .
7. The eukaryotic cell of claim 6 , further comprising one or more additional Cas nucleoprotein complexes.
8. The eukaryotic cell of claim 7 , wherein the eukaryotic cell is a human cell.
9. The Cas nucleoprotein complex of claim 1 , wherein the Cas protein is catalytically active.
10. The Cas nucleoprotein complex of claim 1 , wherein the Cas protein comprises a fusion protein.
11. A Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated (Cas) nucleoprotein complex, wherein the nucleoprotein complex comprises a Cas protein comprising an amino acid sequence selected from the group consisting of SEQ ID NO:37, SEQ ID NO:39, and SEQ ID NO:44 and a cognate nucleic acid guide comprising a repeat sequence and a spacer sequence, wherein the repeat sequence and the spacer sequence do not naturally occur together, wherein the Cas protein is capable of forming the nucleoprotein complex with the cognate nucleic acid guide and wherein the complex is capable of site-directed binding to a target nucleic acid sequence.
12. The Cas nucleoprotein complex of claim 11 , wherein the Cas protein comprises the amino acid sequence of SEQ ID NO: 37.
13. The Cas nucleoprotein complex of claim 11 , wherein the Cas protein comprises the amino acid sequence of SEQ ID NO: 39.
14. The Cas nucleoprotein complex of claim 11 , wherein the Cas protein comprises the amino acid sequence of SEQ ID NO: 44.
15. The Cas nucleoprotein complex of claim 4 , wherein the Cas protein comprises the amino acid sequence of SEQ ID NO: 44.
16. The Cas nucleoprotein complex of claim 1 , wherein the Cas protein is catalytically inactive.