| β-lactoglobulin is the richest protein in lactalbumin.It has many important bioactivities,including antibiosis,antivirus,ability of relieving hypertension and reducing cholesterol content byits corresponding polypeptides formed after enzymatic hydrolysis.β-lactoglobulin is among the family of lipid protein carrier.It is able to combine with many hydrophobic molecules and is biologically compatible and degradable,and thus it could be chosen as an ideal carrier.Perfluorinated compounds(PFCs)are fluorinated organic compounds that are widely used in industry and civil use.PFCs are hydrophobic and oleophobic with high surface activity,high heat and chemical resistance.No photolysis,hydrolysis or biodegradation of PFCs have been observed in current environmental tests.PFCs persist in environment and accumulate in food chain.Perfluorooctanoic(PFOA)and perfluooctane sulphonate(PFOS)are the most widely used PFCs.It is reported that many kinds of PFCs have been detected in breast milk and dairy products,and evidences show that PFOA and PFOS can accumulate in dairy products.The interactions of PFOA and PFOS with protein may affect the normal physiological functions of β-lactoglobulin.In this thesis,ESI-MS is applied to study the interaction of PFOA/ PFOS and β-lactoglobulin.It discoveres that PFOA/PFOS can combine with β-lactoglobulin,and stochiometry of complex molecule increases with the concentration.Binding constant of PFOA/PFOS and β-lactoglobulin has been calculated and the results indicate that the binding ability of PFOS is stronger than that of PFOA.Results of fluorescence spectroscopy and competition experiments confirm that PFOA/PFOS combine on the surface of β-lactoglobulin.Auto Dock(v.4.2)is widely used molecular simulation software.In this thesis,Auto Dock is applied to perform the intermolecular semi-flexible docking,that is the spinning of chemical bonds in complex molecule and the change of conformation of small molecules are allowed.The screening of the best combination method is based on Lamarck genetic algorithm.A cluster is generated after evaluation by scoring function.We choose the conformation with the smallest combination energy from the largest cluster and study the residual amino acides which form hydrogen bonds with ligands.Docking results show that PFOA/ PFOS bind to the surface of β-lactoglobulin.Variant A of β-lactoglobulin binds to PFOA with the binding energy of-3.51 Kcal/mol and binds to PFOS with the energy of-3.79 Kcal/mol.Variant B of β-lactoglobulin binds to PFOA with the energy of-4.59 Kcal/mol and binds to PFOS at energy of-5.41 Kcal/mol.Therefore,the binding ability of PFOS is stronger than that of PFOA.PFOS contains a sulfonic acid group,which can form 3 hydrogen bonds with residual amino acids;while the terminal of PFOA is a carboxyl group,which can only form 2 hydrogen bonds.Formation of hydrogen bonds plays a crucial role in the non-covalent interaction between PFCs and β-lactoglobulin.Flavonoid family,which widely exists in fruit,vegetable,grain,herb and tea,possesses a lot of biological functions such as anti-oxidation,diminishing inflammation,preventing cancer,preventing CVD and controlling virus propagation,and thus is regarded as functional food composition.Up to date,the interaction between flavonoid and β-lactoglobulin has not been reported.Study of interactions between nutrient substances helps to understand the compatibility of foods.In this thesis,we chose β-lactoglobulin as the protein carrier and morin,quercetin,myricetrin as ligands.We studied the interaction between flavonoids and β-lactoglobulin by ESI-MS;optimized ESI conditions for the observation of their interaction.Hydrogen bonds between hydroxyl group in phenol and protein may contribute to the formation of complex molecule between flavonoids and β-lactoglobulin.The order of binding ability of ligands is morin > quercetin > myricetrin... |
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