Detection of Protease Activity by Concentration Quenching-Based Substrates

摘要

Concentration quenching-based substrates have been developed for protease assays [1,2]. The substrate fluorescence is quenched by concentration quenching resulting from highly assembled fluorophores on the substrates. The release of these concentrated fluorophores by proteases leads to fluorescence recovery, thereby allowing detection of protease activity. In this study, we designed and synthesized model compounds as concentration quenching-based substrates. They are composed of two fluorescein isothiocyanates (FITCs) and one or two substrate peptides (|3-Ala-L-Ala-L-Phe-L-Ala) (Figure 1). Two types of concentration quenching-based substrates were designed: short-type substrates and long-type substrates. The short-type substrates are comprised of one FITC-linked substrate peptide and FITC on each edge of the core respectively. The long-type substrates are comprised of two FITC-linked substrate peptides. These substrates include hexamethylenediamine or ethylenediamine as the core.