Study On The Interaction Between Small-molecular Drugs And Bovine Serum Albumin

Small molecular drugs are always inhibitors of signal pathway, which cancombine with target proteins specifically, and then block relative signal pathway,finally obtaining the purpose of treatment. Serum albumin, which is the mostabundant protein of plasma, can combine with numbers of endogenous and exogenoussubstances, and is an important carrier and target for drugs to take effect. Whenentering the blood, drugs will generally bind to serum albumin with varying degrees,which will affect the absorbance, metabolism, efficiency, pharmacological andtoxicological effects of drugs. The knowledge on the interaction between drug andprotein will contribute to the design and modification of drugs, and is also animportant way of understanding the biological effect of proteins, which will be ofgreat significance on pharmacodynamics, pharmacokinetics, pharmacology andtoxicology.So far, it is difficult to determine the binding of drugs and proteins directly invivo, as a result, the determination of their binding is always carried out in vitrosimulated conditions. Bovine serum albumin has become one of the most studiedmodel proteins on drug-protein interaction.Eupatorin and aucubin are two small-molecular drugs with significant drugefficiency and good development prospects. However, study on eupatorin-proteininteraction hasn’t been reported, though there’s few investigations on aucubin-proteininteraction, the interaction mechanism hasn’t been reported, which have become thelimiting factors on development of the two drugs. On the basis of predecessor’s work,this paper has studied the interaction between eupatorin, aucubin and BSAsystematically. The innovative research has mainly focused on the following aspects:1. Under physiological conditions, UV-vis spectroscopy and fluorescencespectroscopy have been used to study the interaction between eupatorin, aucubin and BSA, and the interaction mechanism （fluorescence quenching mechanism, bindingconstants and number of binding sites） has been confirmed, the interaction forceshave been discussed, and the binding distance of eupatorin and BSA has beencalculated.2. Synchronous fluorescence and circular dichroism have been used to study theeffect of eupatorin and aucubin on secondary structure of BSA.3. Molecule docking method has been used to characterize the interactionbetween eupatorin and bovine serum albumin, and its results are in accord with theseof spectral experiments.4. The interaction between aucubin and BSA has been studied with or withoutCu²⁺and Fe³⁺.This paper consists of three parts:PART1: Research progress on small molecule-protein interactionsIt is the introduction of this paper, mainly on the structure and function ofproteins, the effects of metal ions on the interaction between drugs and proteins, andthe significance as well as methodology and main contents on the study of interactionbetween drugs and proteins.PART2: Spectroscopies and molecule docking method to characterize theinteraction between eupatorin and bovine serum albumin.1. UV-vis spectroscopy has been used, and the results show that eupatorin causesthe changes of BSA absorbance at220nm and280nm, which show that eupatorin canbind to bovine serum albumin and lead to the conformational changes of the peptidechain.2. According to the classic and modified Stern-Volmer equations, the results offluorescence spectroscopy have shown that eupatorin results in the fluorescencequenching of BSA. The quenching mechanism has been confirmed to be staticquenching. The binding constant, number of binding sites and interaction forces havebeen confirmed, and the distance to Trp have been calculated by Van’t Hoff equation.3. Synchronous fluorescence has been employed, and the results show that thereis no significant change on the microenvironment of BSA Trp and Tyr residues. The data of CD shows that α-helix content has been reduced significantly resulting fromeupatorin-BSA interaction.4. Molecule docking method has been used to study the interaction betweeneupatorin and bovine serum albumin, and its results are in accord with these ofspectral experiments.PART3: Study the interaction between aucubin and BSA1. Under physiological conditions, the results of fluorescence spectroscopy andStern-Volmer equation have shown that there exists interaction between aucubin andBSA with or without Cu²⁺and Fe³⁺. The presence of aucubin results in the quenchingof BSA, and it is confirmed to be static quenching mechanism in multiple angles, andthe binding constant, number of binding sites and binding forces have been discussedand confirmed. Results of thermodynamic analysis have shown that hydrophobicinteractions play a vital role during the interaction.2. The results of UV spectroscopy have further confirmed that the quenchingmechanism caused by aucubin is static quenching.3. Synchronous fluorescence has been employed in the absence and presence ofCu²⁺and Fe³⁺, and shows that aucubin has no significant effect on themicroenvironment of BSA Trp and Tyr residues for both cases. The data of CD showsthat there is no significant change of α-helix content （slight reduction） onaucubin-BSA interaction.