Study On The Interaction Mechanism And Enzyme Inhibition Kinetics Between Small Drug Molecules Loaded On Nanocarrier And Biopolymers

Currently,drug design to develop natural drugs with active ingredients based on biological macromolecules such as proteins,enzymes and nucleic acids is a hot topic in life science.To study the mechanism of interaction between small drug molecules and biomacromolecules is to explore the pharmacological action of drugs at the molecular level,which is helpful to understand and grasp the operation and metabolism of drugs in vivo,and also provides important information for the further study of pharmacokinetics and pharmacodynamics of drugs in human body,as well as drug screening.The study of the effects of small drug molecules and biomacromolecules has important guiding significance for the design of new effective drugs with low toxicity.It has certain research value and practical significance for the structure of biomolecules,and provides theoretical guidance and valuable information for drug design and development.In this paper,three different nanocomposites were used as drug carriers to study the effect of small drug molecules on the conformation and toxicity of biological macromolecules at the molecular level.The main contents of this thesis are as follows:Chapter 1:The origin and classification of natural plant active ingredients;The reason,basis and significance of biomacromolecules as research objects;Summary of research methods;Overview of nanocomposites and research background as drug carriers.Chapter 2:The interaction between HSA and HCPT supported by MrGO was studied by multispectral method.The experimental results show that the quenching mode of HSA and HCPT is mainly static quenching process,the main driving force is enthalpy,the main force is hydrogen bond and van der Waals force.Although the addition of MrGO did not change the quenching mode,it greatly enhanced the binding effect of HSA and HCPT.In addition,the binding parameters showed that only one binding site existed in the HSA-MrGO-HCPT complex.Combined with fluorescence spectroscopy,infrared spectroscopy and circular dichroism experiments,the conformation of HSA changes slightly regardless of the presence of MrGO.In addition,the comet experiment showed that MrGO-HCPT has less toxic effect on normal somatic cells,indicating that MrGO has good biocompatibility.Chapter 3:ZnO@rGO nanocomposites were synthesized by one-step hydrothermal method.The inhibitory effect and mechanism of ZnO@rGO nanocomposites loaded with curcumin onα-glucosidase（α-Glu）were studied by multispectral method and enzyme inhibition kinetics.The quenching ofα-Glu by ZnO@rGO loaded Cur is a static quenching process with hydrogen bond and van der Waals force,which is mainly driven by enthalpy.What’s more,Tyr residues played a more important role in the fluorescence quenching process.EEM,FT-IR and CD spectra further demonstrated thatα-Glu conformation has changed in the presence of ZnO@rGO and ZnO@rGO-Cur,indicating that ZnO@rGO and ZnO@rGO-Cur both have inhibitory effects onα-Glu.From the data analysis,ZnO@rGO-Cur has a stronger inhibitory effect onα-Glu.Study on the inhibition kinetics ofα-Glu further showed thatα-Glu-Cur belonged to competitive inhibition,α-Glu-ZnO@rGO-Cur belonged to anti-competitive inhibition,and its inhibition constants K_i,Cur>>K_{is,ZnO@rGO-Cur},indicating that ZnO@rGO loaded curcumin had a greater inhibitory effect onα-Glu than pure curcumin.In addition,CCK-8 experiment also proved that ZnO@rGO nanocomposite has good biocompatibility,which indicated that ZnO@rGO is an effective drug nanocarrier loaded with Cur.Chapter 4:Mg-MOF-74 was synthesized by a simple method.Based on the good performance and drug loading effect of Mg-MOF-74,the interaction mechanism and inhibitory mechanism of Mg-MOF-74 loaded quercetin（Que）on pancreatic lipase（PL）were studied by multi-spectral technique and enzyme inhibition kinetics analysis method.Studies show that the effect of pure Que on PL is a static quenching process dominated by hydrogen bond and van der Waals force,while the effect of Mg-MOF-74 loaded with Que on PL is a dynamic quenching process dominated by hydrophobic force.In the quenching process,the binding site was closer to the Tyr residue,in other words,Tyr residue played a more important role in the quenching process.EEM,FT-IR and CD spectra showed that the conformation of PL has changed in different degrees under the influence of Mg-MOF-74 and Mg-MOF-74-Que.Further analysis of the inhibition kinetics of PL shows that PL-Que belongs to mixed inhibition,PL-Mg-MOF-74-Que belongs to uncompetitive inhibition,and the inhibition constants K_i,Que>K_{is,Mg-MOF-74-Que},which indicating that pure Que has less inhibitory effect on PL than Mg-MOF-74-Que.By studying the inhibitory mechanism and interaction mechanism of Mg-MOF-74-Que and PL at the molecular level,it provides a new idea for the design of novel pancreatic lipase inhibitors and a new method for the potential treatment of obesity.