Novel internally quenched substrate of the trypsin-like subunit of 20S eukaryotic proteasome

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• 作者：Natalia Gruba^a ; ¹ ; Magdalena Wysocka^a ; ¹ ; Magdalena Brzezińska^a ; Dawid Debowski^a ; Krzysztof Rolka^a ; Nathaniel I. Martin^b ; Adam Lesner^a ; ^{adam.lesner@ug.edu.pl" class="auth_mail" title="E-mail the corresponding author}
• 关键词：20S Eukaryotic proteasome ; Internally quenched peptides ; Fluorescence
• 刊名：Analytical Biochemistry
• 出版年：2016
• 出版时间：1 September 2016
• 年：2016
• 卷：508
• 期：Complete
• 页码：38-45
• 全文大小：1195 K

文摘

This article describes the synthesis, using combinatorial chemistry, of internally quenched substrates of the trypsin-like subunit of human 20S proteasome. Such substrates were optimized in both the nonprime and prime regions of the peptide chain. Two were selected as the most susceptible for proteasomal proteolysis with excellent kinetic parameters: (i) ABZ-Val-Val-Ser-Arg-Ser-Leu-Gly-Tyr(3-NO₂)-NH₂ (k_cat/K_M = 934,000 M⁻¹ s⁻¹) and (ii) ABZ-Val-Val-Ser-GNF-Ala-Met-Gly-Tyr(3-NO₂)-NH₂ (k_cat/K_M = 1,980,000 M⁻¹ s⁻¹). Both compounds were efficiently hydrolyzed by the 20S proteasome at picomolar concentrations, demonstrating significant selectivity over other proteasome entities.