Patent ID: 6413725
Filing Date: 2002-07-02
Classification: C12Q,Y10S,Y10T

Abstract:
A method of identifying a compound that modulates an activity of a ubiquitin ligase, comprising:(a) contacting a test compound under suitable ubiquitination conditions with a mixture comprising ubiquitin, a ubiquitin-activating E1 enzyme, adenosine tri-phosphate (ATP), yeast ubiquitin-conjugating enzyme Cdc34, a substrate comprising a chimeric protein comprising a ubiquitin conjugating enzyme and a conjugation substrate,1 and an ubiquitin ligase selected from the group consisting of yeast ubiquitin ligase Cdc53, and Cdc53-related human cullins Cul1, Cul2, Cul3, Cul4, Cul5 and Cul6; (b) measuring the ubiquitination of said substrate under the conditions of (a); and (c) comparing the ubiquitination of said substrate as measured in (b) to ubiquitination of said substrate in the mixture when not contacted with said test compound to determine a difference in the ubiquitation of said substrate, wherein the difference is indicative of the ability of said test compound to modulate ubiquitin ligase activity.