A novel fluorogenic probe for the investigation of free thiols: Application to kinetic measurements of acetylcholinesterase activity.
A novel coumarin-derived thiol probe, based on the thiol-promoted cleavage of a quenching 2,4-dinitrobenzenesulfonyl group is described. The probe shows a sensitive fluorescence turn-on and sufficient solubility in aqueous environments. As a proof of concept, a new assay for AChE activity was developed as a useful addition to the established Ellman method. The observed reaction kinetics followed an asymmetric sigmoidal pattern and were successfully evaluated applying a three parameter Gompertz equation. Providing a linear relationship between the detected fluorescence formation curves and corresponding enzyme activities, this probe appears as a valuable tool for AChE activity measurements.
Pharmaceutical Institute, Pharmaceutical Chemistry I, University of Bonn, An der Immenburg 4, 53121 Bonn, Germany.
Matthias D
MD
Mertens
Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 München, Germany.
Anne
A
Bierwisch
Pharmaceutical Institute, Pharmaceutical Chemistry I, University of Bonn, An der Immenburg 4, 53121 Bonn, Germany.
Tianwei
T
Li
Pharmaceutical Institute, Pharmaceutical Chemistry I, University of Bonn, An der Immenburg 4, 53121 Bonn, Germany.
Michael
M
Gütschow
Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 München, Germany.
Horst
H
Thiermann
Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 München, Germany.
Timo
T
Wille
Pharmaceutical Institute, Pharmaceutical Chemistry I, University of Bonn, An der Immenburg 4, 53121 Bonn, Germany; Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 München, Germany. Electronic address: paul.elsinghorst@uni-bonn.de.
Paul W
PW
Elsinghorst
D000110:Acetylcholinesterase / Q000378:metabolism*; D000465:Algorithms; D003374:Coumarins / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000378:metabolism*; D058851:GPI-Linked Proteins / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D016018:Least-Squares Analysis; D017711:Nonlinear Dynamics; D012815:Signal Processing, Computer-Assisted; D013050:Spectrometry, Fluorescence; D013438:Sulfhydryl Compounds / Q000378:metabolism*
D003374:Coumarins; D005456:Fluorescent Dyes; D058851:GPI-Linked Proteins; D013438:Sulfhydryl Compounds; C548244:ACHE protein, human; D000110:Acetylcholinesterase
Acetylcholinesterase; Activity assay; Fluorescence; Thiols; Turn-on probe