Real-time fluorescence assay for monitoring transglutaminase activity

Transglutaminases (TGs) form a family of enzymes that catalyze various posttranslational protein modifications such as crosslinking, esterification and deamidation in a Ca2+-dependent manner.(1) Their main function is the formation of covalent Nε-(γ-glutamyl)lysine bonds within or between polypeptides to stabilize protein assemblies. The activity of these enzymes is crucial for tissue homeostasis and function in a number of organ systems, and the lack of or the excessive crosslinking activity have been linked to human disease processes(1,2). Here we perform kinetic measurements using recombinant TG2 and a fluorescent peptide model substrate on a FLUOstar OPTIMA and FLUOstar Omega in a format suitable for high-throughput analysis. This assay principle can be applied to kinetic studies on closely related enzymes including TG6(3) and can be optimised by modification of the backbone peptide sequence