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US 10,590,462 B2Granted
Probes and assays for measuring E3 ligase activity
AssigneeNorthwestern University
InventorsAlexander V. Statsyuk, Sungjin Park
Life of this patent
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Abstract
Provided herein is technology relating to the biological process of protein ubiquitination and particularly, but not exclusively, to compositions and methods for studying protein ubiquitination and developing therapeutics to modulate protein ubiquitination.
Claims14
A method of screening for modulators of E3 ligase activity, the method comprising:
- a) providing a test composition comprising an E3 ligase, a ubiquitin C-terminal thioester fluorophore, and a molecule to test for the ability to modulate the activity of the E3 ligase;
- b) monitoring the fluorescence of the fluorophore in the test composition;
- c) comparing the fluorescence of the fluorophore in the test composition to the fluorescence of a control composition comprising the E3 ligase and the ubiquitin C-terminal thioester fluorophore, but not comprising the molecule to test;
- and
- d) identifying the molecule to test as a molecule that modulates the activity of E3 ligase when the fluorescence of the fluorophore in the test composition is different than the fluorescence of the fluorophore in the control composition.
2 more elements
A method of measuring the activity of an E3 and/or an E2 enzyme, the method comprising:
- a) providing a test composition comprising an E3 and/or an E2 enzyme and a ubiquitin C-terminal thioester fluorophore;
- b) monitoring the fluorescence of the fluorophore in the test composition;
- and
- c) calculating the activity of the E3 and/or E2 enzyme using the fluorescence of the fluorophore as a function of time.
1 more element
Dependencies are inferred from the claim text.