Identification of Protein Targets of Bioactive Small Molecules Using Randomly Photomodified Probes
| Autor: | Tomáš Knedlík, Vladimir Subr, Pavel Šácha, Anna Březinová, Libor Kostka, Kristýna Blažková, Petra Dvořáková, Jan Konvalinka, Petr Šimon |
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| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Glutamate Carboxypeptidase II Proteomics Fluorophore Ultraviolet Rays Biotin Conjugated system 01 natural sciences Biochemistry Mass Spectrometry 03 medical and health sciences chemistry.chemical_compound Binding efficiency Polymethacrylic Acids Cell Line Tumor Endopeptidases Aspartic Acid Endopeptidases Humans Enzyme Inhibitors Fluorescent Dyes chemistry.chemical_classification Microscopy Confocal 010405 organic chemistry Serine Endopeptidases Membrane Proteins Affinity Labels General Medicine Fluoresceins Combinatorial chemistry Small molecule 0104 chemical sciences 030104 developmental biology Enzyme chemistry Diazomethane Gelatinases Molecular Medicine Identification (biology) Linker |
| Zdroj: | ACS chemical biology. 13(12) |
| ISSN: | 1554-8937 |
| Popis: | Identifying protein targets of bioactive small molecules often requires complex, lengthy development of affinity probes. We present a method for stochastic modification of small molecules of interest with a photoactivatable phenyldiazirine linker. The resulting isomeric mixture is conjugated to a hydrophilic copolymer decorated with biotin and a fluorophore. We validated this approach using known inhibitors of several medicinally relevant enzymes. At least a portion of the stochastic derivatives retained their binding to the target, enabling target visualization, isolation, and identification. Moreover, the mix of stochastic probes could be separated into fractions and tested for binding affinity. The structure of the active probe could be determined and the probe resynthesized to improve binding efficiency. Our approach can thus enable rapid target isolation, identification, and visualization, while providing information required for subsequent synthesis of an optimized probe. |
| Databáze: | OpenAIRE |
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