| Investigation on the interaction between small molecules (ions) and biological macromolecules is a path to explore the biology effects and functions of small molecules (ions), which is helpful to understand life process from molecular level.To understand the interactions between small molecules (ions) and proteins thoroughly, fluorescence spectrum (FS), ultra-violet spectrum (UV) was used to study the binding site of a traditional Chinese Herb active component, berberine chloride (BC), on the pepsin in this paper, and nuclear magnetic resonance (1H NMR) was used to investigate the key segments of BC, sodium benzoate, glucose, Vitamin C(VC) binding bovine serum albumin (BSA) and pepsin , as well as that of imidazole ionic liquids binding BSA. In order to explore the latent toxicity of ionic liquid, multi-spectrum methods were used to study the interaction between imidazole ionic liquids and BSA. Considering the complexity of the interactions among small molecules (ions) binding proteins, the interference of coexisting substances in esculetin-BSA, BC-BSA binding process and the allosteric effects of Cu2+, Zn2+, BC-BSA binding process were studied, the mechanisms were also discussed combined the results of CD spectra. To make the drug-protein interaction parameters more accurate, the interaction parameters between BC and tryptophan, tyrosine were abtained respectively. Generally mathematics rule in structure-performance relationship of drug molecules-serum albumin interaction was also discussed with the model formed by SAS. In the experimental range of this paper, the main conclusions are listed as follow:1. The allosteric effect of BC on Cu2+ quenching BSA intrinsic fluorescence is negative, while the allosteric effect of BC on the stability of Cu2+-BSA complex and the binding sites of Cu2+ is positive. The allosteric effects of Cu2+, Zn2+ on BC quenching BSA intrinsic fluorescence is positive, while the allosteric effects of Cu2+, Zn2+ on the stability of BC-BSA complex and the binding sites of BC is positive.2. BC can insert the hydrophobic cavity of pepsin in special direction, and the average spatial distance between BC and tyrosine is smallest. The key segment of BC binding BSA and pepsin is the group of isoquinoline and benzene ring with conjugateπsystem, its binding orientations existence difference. The key segments of sodium benzoate (SB), glucose, VC binding BSA and the pepsin are different, while their binding site on BSA and pepsin are possibly located in the hydrophilic region on the protein molecular surface.3. Both esculetin-BSA and BC-pepsin binding process are interfered by each selected coexisting substances more or less. BSA molecular conformational transition due to coexisting substances bound to BSA is the common mechanism of coexisting substances participating in esculetin-BSA binding process. The interference mechanism of the selected coexisting substances is different among each other, such as ionic bridge (I-), appositional replacement (SB), destroying the binding sites (SDS), inducing the conformation of BSA changed (TW-80, VC, glucose).4. The key segment of imidazole ionic liquids binding BSA is its cationic part, while the cationic part is not the key group that makes BSA intrinsic fluorescence quenching, and the binding site of imidazole ionic liquids on BSA is not deep, the interaction mechanism is quite complex. Imidazole ionic liquids make BSA the secondary structure change obviously.5. The type of pepsin intrinsic fluorescence being quenched is mainly static quenching but with the dynamic character, the quenching extend of tryptophan is greatest. The BC-pepsin binding is strong, the binding extend of tryptophan is greatest.6. The mathematic rule in structure-performance relationship of drug molecules-serum albumin interaction is not a strict linear relationship. Thus, liminations consist in simply inducing structure-performance relationship of drug molecules-serum albumin interaction based on several specific parameters characterizing the structure of drug molecules.
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