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 having at least 95 percent sequence identity to the amino acid sequence of SEQ ID NO: 40; 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 cognate nucleic acid guide comprises a modified base analog.
5. A eukaryotic cell comprising the Cas nucleoprotein complex of claim 1 .
6. The eukaryotic cell of claim 5 , further comprising one or more additional Cas nucleoprotein complexes.
7. The eukaryotic cell of claim 6 , wherein the eukaryotic cell is a human cell.
8. The Cas nucleoprotein complex of claim 1 , wherein the Cas protein is catalytically active.
9. The Cas nucleoprotein complex of claim 1 , wherein the Cas protein is catalytically inactive.
10. The Cas nucleoprotein complex of claim 1 , wherein the Cas protein comprises a fusion protein.
11. A method of directing a Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated (Cas) nucleoprotein complex to a selected nucleic acid target sequence, the method comprising:
contacting the selected nucleic acid target sequence with one or more nucleoprotein complexes comprising:
a Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated (Cas) protein, wherein the Cas protein has at least 95 percent sequence identity to the amino acid sequence of SEQ ID NO: 40; 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 binds to the selected nucleic acid target sequence.
12. The method of claim 11 , wherein the nucleic acid target sequence comprises RNA.
13. The method of claim 11 , wherein the method is performed in a cell.