Monitoring enzyme reaction and screening of inhibitors of acetylcholinesterase by quantitative matrix-assisted laser desorption/ionization Fourier transform mass spectrometry.
A matrix-assisted laser desorption/ionization Fourier transform mass spectrometry (MALDI-FTMS)-based assay was developed for kinetic measurements and inhibitor screening of acetylcholinesterase. Here, FTMS coupled to MALDI was applied to quantitative analysis of choline using the ratio of choline/acetylcholine without the use of additional internal standard, which simplified the experiment. The Michaelis constant (K(m)) of acetylcholinesterase (AChE) was determined to be 73.9 micromol L(-1) by this approach. For Huperzine A, the linear mixed inhibition of AChE reflected the presence of competitive and noncompetitive components. The half maximal inhibitory concentration (IC(50)) value of galantamine obtained for AChE was 2.39 micromol L(-1). Inhibitory potentials of Rhizoma Coptidis extracts were identified with the present method. In light of the results the referred extracts as a whole showed inhibitory action against AChE. The use of high-resolution FTMS largely eliminated the interference with the determination of ACh and Ch, produced by the low-mass compounds of chemical libraries for inhibitor screening. The excellent correlation with the reported kinetic parameters confirms that the MS-based assay is both accurate and precise for determining kinetic constants and for identifying enzyme inhibitors. The obvious advantages were demonstrated for quantitative analysis and also high-throughput characterization. This study offers a perspective into the utility of MALDI-FTMS as an alternate quantitative tool for inhibitor screening of AChE.
Xu
Zhe
Z
Shanghai Mass Spectrometry Center, Shanghai Institute of Organic Chemistry, Chinese Academy of Science, Shanghai, China.
Yao
Shengjun
S
Wei
Yuanlong
Y
Zhou
Jing
J
Zhang
Li
L
Wang
Cuihong
C
Guo
Yinlong
Y
D000110:Acetylcholinesterase / Q000378:metabolism*; D002800:Cholinesterase Inhibitors / Q000494:pharmacology*; D004353:Drug Evaluation, Preclinical; D005583:Fourier Analysis; D007700:Kinetics; D007834:Lasers; D010936:Plant Extracts / Q000494:pharmacology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D002800:Cholinesterase Inhibitors; D010936:Plant Extracts; D000110:Acetylcholinesterase
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