Assay for kinases and phosphatases
AssigneePromega Corporation
InventorsSaid Goueli, Jolanta Vidugiriene, Natasha Karassina
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Abstract
Disclosed is a method and corresponding kit for assaying the presence, activity, or both, of an enzyme classified within an enzyme classification selected from the group consisting of EC 2.7.1, EC 3.1.3, and EC 3.1.4. The method generally includes the steps of reacting an enzyme with a substrate for a time sufficient to yield phosphorylated or dephosphorylated product; contacting the product with a binding matrix, whereby product is adhered to the matrix; and then analyzing the matrix for presence of, amount of, or both the presence and the amount of the product fixed to the matrix, whereby the presence, the activity, or both the presence and activity of the enzyme can be determined.
Claims16
A method for assaying presence, activity, or both, of an enzyme, the method comprising:
- (a) contacting an enzyme substrate specifically reactive with an enzyme selected from the group consisting of enzymes that catalyze transfer of a phosphate group to a phospholipid substrate, and enzymes that catalyze removal of a phosphate group from a phospholipid substrate;
- whereby the enzyme substrate is fixed to the matrix;
- and then
- (b) contacting the substrate fixed to the matrix with an enzyme under conditions wherein the enzyme is active for a time sufficient to yield phosphorylated product fixed to the matrix when assaying a lipid or phospholipid kinase or a dephosphorylated product fixed to the matrix when assaying a lipid or phospholipid phosphatase;
- and then
- (c) analyzing the matrix for presence of, amount of, or both the presence and the amount of the product fixed to the matrix, whereby the presence, the activity, or both the presence and activity of the enzyme can be determined.
Dependencies are inferred from the claim text.