Autoren:	Maus, Hannah Müller, Patrick Meta, Mergim Hoba, Sabrina N. Hammerschmidt, Stefan J. Zimmermann, Robert A. Zimmer, Collin Fuchs, Natalie Schirmeister, Tanja Barthels, Fabian
Titel:	Next generation of fluorometric protease assays : 7-nitrobenz-2-oxa-1,3-diazol-4-yl-amides (NBD-amides) as class-spanning protease substrates
Online-Publikationsdatum:	14-Sep-2023
Erscheinungsdatum:	2023
Sprache des Dokuments:	Englisch
Zusammenfassung/Abstract:	Fluorometric assays are one of the most frequently used methods in medicinal chemistry. Over the last 50 years, the reporter molecules for the detection of protease activity have evolved from first-generation colorimetric p-nitroanilides, through FRET substrates, and 7-amino-4-methyl coumarin (AMC)-based substrates. The aim of further substrate development is to increase sensitivity and reduce vulnerability to assay interferences. Herein, we describe a new generation of substrates for protease assays based on 7-nitrobenz-2-oxa-1,3-diazol-4-yl-amides (NBD-amides). In this study, we synthesized and tested substrates for 10 different proteases from the serine-, cysteine-, and metalloprotease classes. Enzyme- and substrate-specific parameters as well as the inhibitory activity of literature-known inhibitors confirmed their suitability for application in fluorometric assays. Hence, we were able to present NBD-based alternatives for common protease substrates. In conclusion, these NBD substrates are not only less susceptible to common assay interference, but they are also able to replace FRET-based substrates with the requirement of a prime site amino acid residue.
DDC-Sachgruppe:	540 Chemie 540 Chemistry and allied sciences
Veröffentlichende Institution:	Johannes Gutenberg-Universität Mainz
Organisationseinheit:	FB 09 Chemie, Pharmazie u. Geowissensch.
Veröffentlichungsort:	Mainz
ROR:	https://ror.org/023b0x485
DOI:	http://doi.org/10.25358/openscience-9514
Version:	Published version
Publikationstyp:	Zeitschriftenaufsatz
Nutzungsrechte:	CC BY
Informationen zu den Nutzungsrechten:	https://creativecommons.org/licenses/by/4.0/
Zeitschrift:	Chemistry - a European journal 29 50
Seitenzahl oder Artikelnummer:	e202301855
Verlag:	Wiley-VCH
Verlagsort:	Weinheim
Erscheinungsdatum:	2023
ISSN:	1521-3765
DOI der Originalveröffentlichung:	10.1002/chem.202301855
Enthalten in den Sammlungen:	DFG-491381577-H