Interactions Of Five Natural Flavonoids With CYP3A4 And CYP2D6

The fluorescence quenching mechanism of five natural flavonoids,namely astilbin,neoastilbin,isoastilbin,engeletin,coreopsin,on CYP3A4/CYP2D6 was studied by fluorescence spectroscopy.The experimental results showed that the quenching mode of coreopsin on the two enzymes was mainly static quenching,but also accompanied by some dynamic quenching.The quenching mechanism of the other four flavonoids on the two enzymes was static quenching.The binding capacity of five flavonoids with CYP3A4 was greater than that with CYP2D6.The binding capacity of astilbin to two CYP enzymes was the largest,followed by isoastilbin,neoastilbin,coreopsin and engeletin.According to thermodynamic parameters,it was determined that the binding process of five flavonoids with two CYP enzymes was spontaneous and exothermic,and its main forces were hydrogen bonding and van der Waals force.The energy transfer experiment proved that five flavonoids effectively quenched the endogenous fluorescence of two CYP enzymes through non-radiation energy transfer.The results of Ultraviolet-visible(UV-vis)showed that five flavonoids interacted with two CYP enzymes and formed complexes,which led to the conformational changes of the two CYP enzymes.Synchronous fluorescence and three-dimensional fluorescence(3D)spectroscopy proved that the combination of astilbin,isoastilbin,neoastilbin,engeletin and coreopsin with two CYPs enzymes led to the change of the microenvironment of tyrosine(Tyr)and tryptophan(Trp)residues.Fourier-transform infrared(FT-IR)analysis showed that five flavonoids changed the conformation of the proteins by changing the amide band I and A band of two CYPs enzymes.The circular dichroism(CD)spectrum was used to analyze the changes of the secondary structure such as α-helix,β-sheet and random-coil in the protein before and after the combination of five flavonoids with two CYP enzymes,and it was determined that the five flavonoids mainly changed the conformation of the protein by changing α-helix.The interaction between five flavonoids and two CYP enzymes was simulated by molecular docking,and the conformation with the lowest energy among the most conformations was selected as the optimal binding mode.The binding sites,key amino acids and binding forces of five flavonoids with two CYP enzymes were determined.It was confirmed that five flavonoids were bound to two CYP enzymes mainly through hydrogen bonds and van der Waals forces.