Study On Interaction Mechanism And Structure-Activity Relationship Of Lysozyme With The Active Ingredients In Three Types Of Traditional Chinese Medicine

Objective The interaction mechanism and structure-activity relationship of lysozyme（LYSO） with four furanocoumarin drugs: psoralen, isopsoralen,5-methoxypsoralen,8-methoxypsoralen and two alkaloid drugs: sophocarpine, oxysophocarpine and threehydroxyanthraquinone drugs:1,2-dihydroxyanthraquinone;1,4-dihydroxyanthraquinone,1,8-dihydroxyanthraquinone were investigated under the simulativehuman physiological conditions. The effect of four metal ions and four rare earth ionson the interaction between LYSO and the active ingredient in the three types oftraditional Chinese medicine were explored.Methods The changes of the UV absorbance before and after the interactionbetween the active ingredients in three types of traditional Chinese medicine andLYSO were studied by the ultraviolet difference spectra. Their interactionmechanism, correlating parameters and the interaction force were obtained by thefluorescence spectroscopy. The effect of active ingredient in three types oftraditional Chinese medicine on the conformation of LYSO were studied by thesynchronous fluorescence spectra. Furthermore, the changes of secondary structureof LYSO before and after the interaction with psoralen, isopsoralen,5-methoxypsoralen and8-methoxypsoralen were studied by the circular dichroism.The influence of four metal ions Fe³⁺, Cu²⁺, Zn²⁺, Mg²⁺and four rare earth ions La³⁺,Gd³⁺, Yb³⁺, Lu³⁺on the interaction between the active ingredient in three types oftraditional Chinese medicine and LYSO were investigated.Results The result of ultraviolet difference spectra initially revealed that the active ingredient in three types of traditional Chinese medicine could interact with LYSO.Fluorescence spectra showed that the intrinsic fluorescence of LYSO had beensignificantly quenched by the active ingredient in three types of traditional Chinesemedicine, and the mechanism of fluorescence quenching was static quenching withnon-radiation energy transfer. The major driving forces were hydrogen bond and Vander Waals. The1:1complex was formed between the active ingredient in three typesof traditional Chinese medicine and LYSO. The binding parameters of the activeingredient in three types of traditional Chinese medicine and LYSO at310^K were asfollows: the binding constants（K） were1.02×10⁴,6.95×10⁴,7.05×10⁴,8.60×10⁴,9.22×10⁴,2.81×10⁵,5.51×10⁵,2.01×10⁵and4.05×10⁵L·mol^-1; the binding distance（r）were4.36,4.75,5.25,5.89,3.81,4.89,4.66,5.15and5.69nm, respectively. Theresult of circular dichroism indicated that four furanocoumarin drugs had an impacton the secondary structure of LYSO. The content of α-helical structure of LYSOwas reduced27.1%,11.5%,10^.6%and0.917%by psoralen, isopsoralen,5-methoxypsoralen and8-methoxypsoralen, respectively. The result of synchronousfluorescence spectra demonstrated that the conformation of LYSO was changeddifferently by the active ingredient in three types of traditional Chinese medicine. Theinfluence of four metal ions Fe³⁺, Cu²⁺, Zn²⁺, Mg²⁺and four rare earth ions La³⁺,Gd³⁺,Yb³⁺,Lu³⁺on the interaction of the active ingredient in three types of traditionalChinese medicine with LYSO were different respectively. It is speculated that fourmetal ions such as Fe³⁺participate in the active ingredient in three types of traditionalChinese medicine and LYSO interaction process by means of “ionic bridge” mode.The four rare earth ions participated in the process of active ingredients in three typesof traditional Chinese medicine and LYSO in different modes. It is conjectured thatLa³⁺and Gd³⁺participate in the three types of traditional Chinese medicine-LYSOinteraction process by means of “ionic bridge” mode; Yb³⁺and Lu³⁺participate in the three types of traditional Chinese medicine-LYSO interaction process by means of“appositional substitution”mode.Conclusion The active ingredient in three types of traditional Chinese medicinecould interact with LYSO. The interactions between the active ingredient in threetypes of traditional Chinese medicine and LYSO were influenced differently by fourmetal ions Fe³⁺, Cu²⁺, Zn²⁺, Mg²⁺and four rare earth ions La³⁺, Gd³⁺,Yb³⁺,Lu³⁺,respectively.