Tissue-restricted bioreactions can be utilized to design chemical-biological tools and prodrugs. We have developed a fluorescent-substrate-library-based enzyme discovery approach to screen tissue extracts for enzymatic activities of interest. Assay-positive candidate proteins were identified by diced electrophoresis gel assay followed by peptide mass fingerprinting. We discovered that pyruvyl anilide is specifically hydrolyzed by carboxylesterase 2 (CES2), which is predominantly localized in the liver and kidney. We show that the pyruvyl targeting group/CES2 enzyme pair can be used to deliver the 7-amino-4-methylcoumarin fluorophore specifically to the liver and kidney in vivo. Our screening approach should be useful to find other masking group/enzyme pairs suitable for development of fluorescent substrates and prodrugs.
Yoshioka K, Komatsu T, Nakada A, Onagi J, Kuriki Y, Kawaguchi M, Terai T, Ueno T, Hanaoka K, Nagano T, Urano Y (2015) Identification of Tissue-Restricted Bioreaction Suitable for in Vivo Targeting by Fluorescent Substrate Library-Based Enzyme Discovery Journal of the American Chemical Society137: 12187-90
Yoshioka K, Komatsu T, Nakada A, Onagi J, Kuriki Y, Kawaguchi M, Terai T, Ueno T, Hanaoka K, Nagano T, Urano Y (2015) Journal of the American Chemical Society137: 12187-90