Evaluation method for compatibility of compounds in fluorescence resonance energy transfer model to screen β-secretase inhibitors
10.3867/j.issn.1000-3002.2018.05.004
- VernacularTitle:基于荧光共振能量转移法建立筛选β分泌酶抑制剂的适用性评价方法
- Author:
Ying ZHAO
1
;
Jia-Huai ZHANG
Author Information
1. 中国医学科学院北京协和医学院药物研究所
- Keywords:
fluorescence resonance energy transfer;
β-secretase;
fluorescence intensity;
(-)-epigallocatechin-3-gallate
- From:
Chinese Journal of Pharmacology and Toxicology
2018;32(5):371-376
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE To develop a method to evaluate the compatibility of compounds in the fluorescence resonance energy transfer (FRET) model for β-secretase (BACE1) inhibitor screening.METHODS Two commercially available BACE1 inhibitor screening systems based on FRET were selected to evaluate the BACE1 inhibitory activities of (-)-epigallocatechin-3-gallate (EGCG) and Compound 1 according to the supplier's protocol.The inhibitory rates and slopes of the catalytic curves of the inhibitors were calculated.The effect of inhibitors on the fluorescence intensity of the systems were quantitatively calculated and the comparatively evaluated.RESULTS EGCG,a reported non-competitive inhibitor of BACE1,directly induced the reduction of fluorescence intensity of one of the systems.The slope of the line with the addition of EGCG (10.8±2.6) conformed to that of the line of EGCG inhibition (10.2±3.4),which indicated that EGCG was a pseudo-positive inhibitor of BACE1.Compound 1 had little effect on the fluorescence intensity of the systems,so the inhibitory activity of Compound 1 was confirmed.The compounds which showed inhibitory activity in preliminary screening should be checked in the blank control without BACE1 to calibrate the effect of compound on the system fluorescence intensity.The applicability of the tested compounds in the screening system could thus be evaluated to prevent pseudo-positive results.CONCLUSION This fluorescence calibration method with compound control can be universally used for assays based on FRET theory to evaluate the applicability of tested BACE1 inhibitors.
