Sortases are a family of membrane–anchored transpeptidases expressed by Gram–positive bacteria. Sortase A catalyzes the cleavage of C-terminal recognition motif (LPXTG) of multiple structurally unrelated proteins followed by formation of an amide bond with the peptidoglycan layer of the bacteria. Since sortases are widely distributed among a variety of bacterial pathogens and required for virulence, they represent a promising therapeutic target for the development of novel anti-infective agents. In addition, Sortase A can be used as a biotechnology tool for a variety of protein modifications and immobilization by sortase-mediated protein ligation.
The SensoLyte® 520 Sortase A Assay Kit employs a novel internally quenched 5-FAM/QXL® FRET substrate for the detection of protease activity. The enzyme cleaves the FRET substrate into two separate fragments resulting in the release of 5-FAM fluorescence, which can be monitored at excitation/emission= 490/520 nm. The long wavelength fluorescence of 5-FAM is less interfered by the autofluorescence of components in biological samples and test compounds.