Study On Interaction Of Some Plant Active Components With Protein

In this thesis, the interaction of some plant active components with proteins were studied using different techniques viz., fluorescence, UV-vis absorption, Fourier transform infrared (FT-IR), and circular dichroism (CD) spectroscopy as well as atomic force microscopy (AFM), coupled with molecule docking technique. According to several studies such as intrinsic fluorescence quenching, conformation changes of protein and the effect of adscititious reagents on the interaction between drug and protein, elaborating the binding mechanism in molecular level. This study is helpful to further understand the transportation and metabolic process of the active components from plants in body and provide important information on designing and exploiting new drugs.The thesis consists of five parts and the main conclusions are summarized as follows.1. In this chapter, the structures and functions of BSA, HSA and trypsin were introduced. The development situation and methods of drug molecules and protein interaction were summarized briefly. This thesis was developed based on the apply actuality of fluorescence spectroscopy, Fourier transform infrared spectroscopy and circular dichroism especially in this research field.2. The interaction between farrerol and bovine serum albumin (BSA) in physiological buffer solution (pH7.4) was investigated by fluorescence, UV-vis, Fourier transform infrared and circular dichroism spectroscopy. It was proved from the resuluts that static quenching esits between farrerol and BSA by the formation of BSA-farrerol complexes. The thermodynamic parameters, enthalpy change (△H) and entropy change (△S), were calculated to be-29.92kJ-mol-1and5.06J·mol-1·K-1, respectively, which suggested that the both hydrophobic interactions and hydrogen bonds play major role in the binding of farrerol to BSA. The competitive experiments of site markers suggested that the binding site of farrerol to BSA was sudlow site I. Moreover, the results of FT-IR and CD spectra demonstrated that the microenvironment and the secondary structure of BSA were changed in the presence of farrerol. In addition, the effect of vitamin C (VC), vitamin B2(VB2) or glucose on the interaction between farrerol and BSA were investigated.3. The binding effect of hyperoside on human serum albumin (HSA) was studied by fluorescence, resonance light scattering, CD spectroscopy and atomic force microscopy (AFM), as well as molecule docking. There was a strong fluorescence quenching reaction of hyperoside to HSA, and the interaction between hyperoside and HSA was driven mainly by hydrophobic interactions and hydrogen bonds. Analysis of CD spectra showed that the binding of hyperoside to HSA could induce partial changes in the secondary structure of the protein. The results of AFM and resonance light scattering revealed that the individual HSA molecules was largened and aggregated after interaction with hyperoside. The interaction between hyperoside and HSA was predicted using the molecular docking method:hyperoside might locate in the subdomain IIA (Site I) of HSA, which coincides with the results of displacement experiment of the site markers.4. The interaction between diosmetin and HSA has been studied by using different spectroscopic techniques viz., fluorescence, UV-vis absorption, FT-IR spectroscopy and circular dichroism, as well as AFM. The results showed that the probable quenching mechanism of fluorescence of HSA by hyperoside was a static quenching. The calculated thermodynamic parameters suggested that the binding mode was hydrophobic interactions. It was proved that diomestin was located near the tryptophan residue region of subdomain IIA (sudlow site I) in HSA by competitive binding experiments and unfolding pathways of HSA. The results of FT-IR and CD spectra suggested that the secondary structure of HSA were changed by diomestin. In addition, the results of AFM and resonance light scattering revealed that the individual HSA molecules were aggregated after interaction with diomestin.5. The interaction of mangiferin (MA) with trypsin in pH7.4butter solution has been investigated by fluorescence and CD spectroscopy techniques. The results showed that the quenching mechanism of fluorescence of trypsin by MA was a static quenching by forming the ground-state MA-trypsin complex. The thermodynamic parameters according to vant't Hoff equation showed that the hydrophobic interactions plays major role in the binding of MA to trypsin. The binding locality was an area3.99nm away from tryptophan residue in trypsin based on the Forster theory of non-radioactive energy transfer. Analysis of three-dimensional fluorescence spectra and CD spectra showed that the binding of MA to trypsin could induce partial changes in the secondary structure of the protein.