Proteins that efficiently generate singlet oxygen

Patent metadata

Field	Value
Publication number	US-9279769-B2
Application number	US-201113992540-A
Country	US
Kind code	B2
Filing date	Dec 7, 2011
Priority date	Dec 7, 2010
Publication date	Mar 8, 2016
Grant date	Mar 8, 2016

Abstract

The present invention provides miniSOG proteins, polynucleotides, and methods of use. When expressed in a bacterial or mammalian cell, miniSOG proteins spontaneously incorporate flavin mononucleotide and produce fluorescence and singlet oxygen upon excitation. Uses include optical and electron microscope imaging, in vivo imaging, detection and localization of protein-protein interactions, and photoablation.

First claim

What is claimed is:

1. An isolated polynucleotide encoding a miniSOG polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1 and SEQ ID NO:2.

2. The isolated polynucleotide of claim 1 , wherein the miniSOG comprises an amino acid sequence of SEQ ID NO:1.

3. The isolated polynucleotide of claim 1 , wherein the miniSOG polypeptide consists of 106 amino acids.

4. The isolated polynucleotide of claim 1 , wherein the miniSOG polypeptide has a singlet oxygen quantum yield of 0.47±0.05 or more when bound to a flavin mononucleotide.

5. The isolated polynucleotide of claim 1 , wherein the polynucleotide encoding the miniSOG polypeptide is codon optimized for expression in E. coli or a eukaryotic cell.

6. The isolated polynucleotide of claim 5 , wherein the eukaryotic cell is a mammalian cell.

7. The isolated polynucleotide of claim 5 , wherein the mammalian cell is a human cell.

8. A vector comprising the polynucleotide sequence of claim 1 .

9. A host cell comprising the vector of claim 8 .

10. A miniSOG polypeptide encoded by the polynucleotide of claim 1 .

11. A kit comprising the polynucleotide of claim 1 .

12. A fusion protein comprising a polypeptide encoded by the polynucleotide of claim 1 .

13. A method of expressing in a cell a fusion protein, the method comprising the step of: expressing in a cell a fusion protein comprising i)a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1 and SEQ ID NO:2 and ii) a protein if interest.

14. The isolated polynucleotide of claim 1 , wherein the miniSOG comprises an amino acid sequence of SEQ ID NO:2.

Assignees

• Shu Xiaokun
• Tsien Roger Y
• Univ California

Inventors

• Shu Xiaokun
• Tsien Roger Y

Classifications

• C12N9/12 Primary

  transferring phosphorus containing groups, e.g. kinases (2.7) · CPC title

• G01N21/6486 Primary

  Measuring fluorescence of biological material, e.g. DNA, RNA, cells (G01N21/6428 takes precedence) · CPC title

• C07K14/001

  by chemical synthesis · CPC title

• C12Y207/11001

  Non-specific serine/threonine protein kinase (2.7.11.1), i.e. casein kinase or checkpoint kinase · CPC title

• C07K2319/00

  Fusion polypeptide · CPC title