Opsin polypeptides and methods of use thereof
View Patent ↗The present disclosure provides opsins, including variant opsins with increased activity and/or increased trafficking to the plasma membrane. The opsins are useful in therapeutic and screening applications, which are also provided.
1. An isolated fusion polypeptide comprising an amino acid sequence having at least 85% amino acid sequence identity to the amino acid sequence set forth in SEQ ID NO: 22, wherein said polypeptide is fused to an endoplasmic reticulum (ER) export sequence and/or a trafficking sequence.
2. An isolated polynucleotide comprising a nucleotide sequence encoding an isolated fusion polypeptide comprising an amino acid sequence having at least 85% amino acid sequence identity to the amino acid sequence set forth in SEQ ID NO: 22, wherein said polypeptide is fused to an endoplasmic reticulum (ER) export sequence and/or a trafficking sequence.
3. A recombinant vector comprising the polynucleotide of claim 2 .
4. A genetically modified cell comprising the recombinant vector of claim 3 .
5. The polypeptide of claim 1 , comprising an amino acid sequence having at least about 90% amino acid sequence identity to the amino acid sequence set forth in SEQ ID NO:22.
6. The polypeptide of claim 1 , comprising an amino acid sequence having at least about 95% amino acid sequence identity to the amino acid sequence set forth in SEQ ID NO:22.
7. The polypeptide of claim 1 , wherein the ER export sequence comprises the amino acid sequence VKESL (SEQ ID NO: 1), VLGSL(SEQ ID NO: 2), or FCYENE (SEQ ID NO: 5).
8. The polypeptide of claim 1 , wherein the trafficking sequence comprises the amino acid sequence KSRITSEGEYIPLDQIDINV (SEQ ID NO:10).
9. The polynucleotide of claim 2 , wherein the polynucleotide is codon optimized for expression in a mammalian cell.
10. The polynucleotide of claim 2 , wherein the nucleotide sequence is operably linked to a promoter.
11. The polynucleotide of claim 2 , wherein the nucleotide sequence is operably linked to a neuron-specific transcription control element.
12. The recombinant vector of claim 3 , wherein the recombinant vector is a lentiviral vector, an adenoviral vector, or an adeno-associated viral vector.
13. The genetically modified cell of claim 4 , wherein the cell is a mammalian cell.
14. The genetically modified cell of claim 13 , wherein the cell is a neuron.