Please use this identifier to cite or link to this item:
http://doi.org/10.25358/openscience-9514
Authors: | Maus, Hannah Müller, Patrick Meta, Mergim Hoba, Sabrina N. Hammerschmidt, Stefan J. Zimmermann, Robert A. Zimmer, Collin Fuchs, Natalie Schirmeister, Tanja Barthels, Fabian |
Title: | Next generation of fluorometric protease assays : 7-nitrobenz-2-oxa-1,3-diazol-4-yl-amides (NBD-amides) as class-spanning protease substrates |
Online publication date: | 14-Sep-2023 |
Year of first publication: | 2023 |
Language: | english |
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: | 540 Chemie 540 Chemistry and allied sciences |
Institution: | Johannes Gutenberg-Universität Mainz |
Department: | FB 09 Chemie, Pharmazie u. Geowissensch. |
Place: | Mainz |
ROR: | https://ror.org/023b0x485 |
DOI: | http://doi.org/10.25358/openscience-9514 |
Version: | Published version |
Publication type: | Zeitschriftenaufsatz |
License: | CC BY |
Information on rights of use: | https://creativecommons.org/licenses/by/4.0/ |
Journal: | Chemistry - a European journal 29 50 |
Pages or article number: | e202301855 |
Publisher: | Wiley-VCH |
Publisher place: | Weinheim |
Issue date: | 2023 |
ISSN: | 1521-3765 |
Publisher DOI: | 10.1002/chem.202301855 |
Appears in collections: | DFG-491381577-H |
Files in This Item:
File | Description | Size | Format | ||
---|---|---|---|---|---|
![]() | next_generation_of_fluorometr-20230914114142028.pdf | 3.24 MB | Adobe PDF | View/Open |