Polypeptide immobilization
AssigneeThe University of Chicago
InventorsMilan Mrksich, Christian Hodneland
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Abstract
A substrate comprises a surface, and a plurality of moieties, on at least a portion of the surface. The moieties are moieties of formula: Surf-L-Q-T, where -T comprises a reactant ligand, and Surf- designates where the moiety attaches to the surface. The substrate can be made into a protein chip by the reaction of a reactant ligand and a fusion polypeptide, where the fusion polypeptide includes a capture polypeptide moiety which corresponds to the reactant ligand.
Claims14
A method of assaying kinase activity, comprising:
- contacting a mixture comprising at least one kinase with a protein chip,
- wherein the protein chip comprises
- a substrate with a surface having para-nitrophenyl phosphonate immobilized on the surface;
- and
- covalently immobilized on the surface, a reaction product of a said para-nitrophenyl phosphonate and a fusion polypeptide of cutinase and a kinase substrate;
- and
- correlating a change in the kinase substrate with kinase activity.
A method of assaying protease activity, comprising:
- contacting a mixture comprising at least one protease with a protein chip,
- wherein the protein chip comprises
- a substrate with a surface having para-nitrophenyl phosphonate immobilized on the surface;
- and
- covalently immobilized on the surface, a reaction product of a said para-nitrophenyl phosphonate and a fusion polypeptide of cutinase and a protease substrate;
- and
- correlating a change in the protease substrate with protease activity.
A method of determining the activity of an enzyme in a sample, comprising:
- contacting a protein chip with the sample,
- wherein the protein chip comprises a substrate with a surface having para-nitrophenyl phosphonate immobilized on the surface;
- and covalently immobilized on the surface, a reaction product of said para-nitrophenyl phosphonate and a fusion polypeptide of cutinase and an enzyme substrate specific for the enzyme;
- and
- correlating a change in the enzyme substrate with enzymatic activity.
Dependencies are inferred from the claim text.