Patent ID: 7842460

Claim:
A method for assessing the activity of a proteolytic enzyme, said method comprising: (a) providing a fusion polypeptide comprising: (i) a first polypeptide that comprises Ubiquitin or a Ubiquitin-like protein (UBL), and (ii) a second polypeptide comprising an amino acid sequence having a free N-terminal amino acid required for enzymatic activation and detection, wherein said second polypeptide is an enzyme; wherein the first and second polypeptides are operatively linked to one another through the first polypeptide C-terminus and the second polypeptide N-terminus, and wherein said second polypeptide is enzymatically inactive when operatively linked to said first polypeptide in said fusion polypeptide; (b) contacting the fusion polypeptide with the proteolytic enzyme to produce a cleaved first polypeptide and a cleaved second polypeptide, wherein said cleaved second polypeptide has enzymatic activity; (c) detecting a signal associated with the enzymatic activity of the cleaved second polypeptide; and (d) establishing a correlation between the signal associated with the enzymatic activity of the cleaved second polypeptide and the activity of the proteolytic enzyme, thereby determining the activity of the proteolytic enzyme, wherein the second polypeptide is selected from the group consisting of a 3Dpol RNA-dependent RNA polymerase, GPATase, tryptase, enterokinase, and phospholipase.