Tang, ZH; Gao, JM; Jia, YT; Hu, WX; Hu, JY
To find potential pollutants that had activity effects of LXRs in complex environmental mixtures. In this study, taking the LXRα protein as an example, the pCold-TF-LXRα recombinant protein was constructed using the principle of affinity binding of nuclear receptor protein and small molecules, and the expression conditions of the recombinant protein were optimized, and a method for specifically capturing active substances of LXRα was established. The results showed the optimal expression conditions of recombinant protein when the induction temperature was 20℃ and the concentration of the inducer IPTG was 0.4mmol/L in the protein production system of LXRα. The linear regression curve measured by the spiked recovery experiment of 4gradient concentrations of LXRα agonist T0901317(0.25, 2.5, 25, 250 μg/L) was y = 0.83604x + 0.40763, R2 = 0.9948, proving the effectiveness of the method. Comparing the recovery rate of empty carrier protein which was the pCold-TF (6.42%) and recombinant protein (79.83%) to T0901317 (250μg/L), the method is specific. For proofing the practicality of the method, recombinant protein and a mixed standard sample of 15typical OPEs was subjected to a "capture" experiment. Among them, TPHP, BPADP, TCrP, EHDPP, TNBP, RDP, TEHP, TDCIPP were identified to have the active effect of LXRα. Lastly, the yeast two-hybrid experiment was used to verify that all of active OPEs are LXRα antagonists. © 2020, Editorial Board of China Environmental Science. All right reserved.