Mologic Limited

James Alexander Schouten

Paul James Davis

Gita Parekh

The present invention relates to detecting cleavage activity of an enzyme. The various aspects of the invention include an enzyme detection device, kit, method and use for detecting or measuring the presence in a test sample of the activity of an enzyme capable of cleaving a substrate. The invention also relates to indicator and binding molecules useful for carrying out the invention. The enzyme substrate contains a hidden binding site which is only revealed upon cleavage by the enzyme.

Detection of cleavage activity of an enzyme

A polypeptide comprising a chromogenic amino acid. The chromogenic amino acid is flanked by at least one amino acid to the N and C termini thereof. The amine group of the chromogenic amino acid has a pKa of less than 5. The chromogenic amino acid is capable of reacting with a conjugated aldehyde. The polypeptide comprises a target sequence for a target protease which is capable of cleaving the peptide bond comprising the amino group of the chromogenic amino acid.

Protease detection

A peptide clearing agent is provided for clearance of a conjugate of an enzyme and a binding molecule which binds specifically at a target location from a non-target location in a subject. The peptide clearing agent binds the active site of the enzyme. The peptide also binds to the asialoglycoprotein receptor expressed by hepatic cells to facilitate clearance through the liver. The peptide may be glycosylated to facilitate clearance through the liver by binding to hepatic cells expressing an asialoglycoprotein receptor. Typically, the peptide prevents or inhibits enzyme activity upon binding to the enzyme and is not substantially modified by the enzyme activity. The peptide may be based upon the dipeptide amino-naphthoic acid (ANA)-glutamate (GIu) and may comprise the amino acid sequence serine (Ser)-Alanine (Ala)-amino-naphthoic acid (ANA)-glutamate (GIu). In such cases, the enzyme of interest is typically CPG2.

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Beds, United Kingdom

James Alexander Schouten