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US 11,899,021 B2Granted
Luciferase-based thermal shift assays
AssigneePromega Corporation
InventorsMelanie Dart, Lance P. Encell, Thomas Kirkland, Thomas Machleidt, Matthew Robers, Brock F. Binkowski, Keith Wood, Ce Shi
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Abstract
Provided herein are systems and methods for characterizing target/ligand engagement. In particular, luciferase-labeled polypeptide targets are used to detect or quantify target/ligand engagement (e.g., within a cell or cell lysate).
Claims11
A method to detect an interaction between a ligand and a target protein, comprising the steps of:
- (a) incubating (i) a fusion of a target protein and a peptide tag comprising 70% or greater sequence identity with VSGWRLFKKIS (SEQ ID NO:
- 5) with (ii) a complementary polypeptide comprising 70% or greater sequence identity with SEQ ID NO:
- 6, wherein the peptide tag and the complementary polypeptide are capable of forming a bioluminescent complex:
- (i) in the presence of the ligand to produce a test sample, and
- (ii) in the absence of the ligand, to produce a control sample;
- (b) treating said test and control samples under conditions that cause the target protein to unfold partially or completely unfold;
- (c) measuring signal from the bioluminescent complex in said test and control samples;
- and
- (d) comparing the measurement made in step (c) between the test and control samples, wherein alteration of the signal from said bioluminescent complex in the test sample compared to the control sample indicates the presence of the interaction between the ligand and the target protein.
6 more elements
Dependencies are inferred from the claim text.