Back to results
US 7,060,506 B2Granted
Compositions and methods for monitoring the modification of modification dependent binding partner polypeptides
AssigneeCyclacel Limited
InventorRoger Craig
Life of this patent
Estimating the term…
Abstract
The invention provides methods and compositions for monitoring enzymatic activity as a function of the interaction of modification dependant binding partner polypeptides. Association or dissociation of the binding partner polypeptides is dependant upon the addition or removal of a moiety to or from one or both of the binding partner polypeptides or upon proteolytic digestion of one or both of the binding partner polypeptides by a modifying enzyme.
Claims18
A method for monitoring activity of one or more enzymes comprising the steps of:
- A. mixing:
- (i) one or more tagged binding partner polypeptides;
- (ii) one or more binding partner polypeptides that bind to said one or more tagged binding partner polypeptides of (i);
- and
- (iii) one or more enzymes that add or remove a moiety to or from said one or more binding partner polypeptides or one or more tagged binding partner polypeptides;
- wherein said one or more binding partner polypeptides are immobilized on a solid support;
- and
- wherein said one or more tagged binding partner polypeptides or said one or more binding partner polypeptides comprise one or more sites for the addition or removal of said moiety, wherein addition or removal of said moiety promotes binding of said one or more binding partner polypeptides with the corresponding one or more tagged binding partner polypeptides;
- under conditions which promote binding of said one or more binding partner polypeptides with said one or more tagged binding partners;
- and
- B. detecting binding of said one or more tagged binding partner polypeptides of (i) to said one or more binding partner polypeptides of (ii), wherein the step of detecting binding comprises adding one or more detector molecules comprising a first region that associates with a tag of said one or more tagged binding partner polypeptides and a second region comprising one or more reporter molecules, wherein detection of binding as a result of said mixing is indicative of enzyme activity.
8 more elements
A method of screening for a candidate modulator of enzymatic activity comprising:
- A. mixing:
- (i) one or more tagged binding partner polypeptides comprising a tag;
- (ii) one or more binding partner polypeptides that bind to said one or more tagged binding partner polypeptides of (i);
- and
- (iii) one or more enzymes that adds or removes a moiety to or from said binding partner polypeptide or said one or more tagged binding partner polypeptides;
- wherein said one or more binding partner polypeptides are immobilized on a solid support;
- and
- wherein said one or more tagged binding partner polypeptides or said one or more binding partner polypeptides comprise one or more sites for the addition or removal of said moiety, wherein addition or removal of said moiety promotes binding of said one or more binding partner polypeptides with the corresponding one or more tagged binding partner polypeptides;
- under conditions which promote binding of said one or more binding partner polypeptides and said one or more tagged binding partner polypeptides;
- and
- B. detecting binding of said one or more binding partner polypeptides to said one or more tagged binding partner polypeptides in both the presence and absence of a candidate modulator of enzymatic activity, wherein the step of detecting binding comprises adding one or more detector molecules comprising a first region that associates with said tag of said one or more tagged binding partner polypeptides and a second region comprising one or more reporter molecules, wherein detection of an amount of binding in the presence of the candidate modulator that is lesser or greater as compared to the amount of binding in the absence of the candidate modulator indicates modulation of enzymatic activity by said candidate modulator.
8 more elements
Dependencies are inferred from the claim text.