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US 2017/0370916 A1Published application
High-sensitive fluorescent energy transfer assay using fluorescent amino acids and fluorescent proteins
AssigneeThe Regents of the University of California
InventorsJiayu LIAO, Yang SONG, Yongfeng ZHAO, Yan LIU
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Abstract
The disclosure provides method and composition utilizing fluorescent amino acids and fluorescent proteins comprising a moiety capable of undergoing FRET. The methods and compositions of the disclosure are useful in analyzing protein structure and function, and screening molecular inhibitors.
Claims21
1-24. (canceled)
A method of performing Förster resonance energy transfer (FRET)-based high throughput screening (HTS), the method comprising:
- a) providing a sample in a plurality of wells of a multi-well plate, wherein the sample comprises a first polypeptide comprising a CyPet moiety and a second polypeptide comprising a YPet moiety;
- b) contacting the sample with a library of candidate agents;
- c) exposing the plurality of wells to a first wavelength of light, wherein the first wavelength of light is suitable for exciting the CyPet moiety;
- d) measuring a second wavelength of light in the plurality of wells, wherein the second wavelength of light is emitted from the YPet moiety;
- and
- e) calculating the ratiometric FRET signal between the CyPet moiety and the YPet moiety in each of the plurality of wells.
3 more elements
A kit comprising:
- a) a multi-well plate;
- b) a first polypeptide comprising a CyPet moiety;
- and
- c) a second polypeptide comprising a YPet moiety and a polypeptide selected from the group consisting of a SUMO ligase, a SUMO-conjugating enzyme, and a SUMO-specific peptidase.
1 more element
Dependencies are inferred from the claim text.