Identification of Tissue-Restricted Bioreaction Suitable for in Vivo Targeting by Fluorescent Substrate Library-Based Enzyme Discovery

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 dice...

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Bibliographic Details
Published in:Journal of the American Chemical Society 2015-09, Vol.137 (38), p.12187-12190
Main Authors: Yoshioka, Kentaro, Komatsu, Toru, Nakada, Akihiro, Onagi, Jun, Kuriki, Yugo, Kawaguchi, Mitsuyasu, Terai, Takuya, Ueno, Tasuku, Hanaoka, Kenjiro, Nagano, Tetsuo, Urano, Yasuteru
Format: Article
Language:English
Subjects:
Anilides - chemistry
Anilides - metabolism
Carboxylesterase - metabolism
Coumarins - chemistry
Coumarins - metabolism
Enzyme Activation
Fluorescence
Humans
Kidney - enzymology
Kidney - metabolism
Liver - enzymology
Liver - metabolism
Substrate Specificity
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Summary: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.
ISSN:0002-7863
1520-5126
DOI:10.1021/jacs.5b05884