Molecular Simulation Of Human Serum Albumin-Ligand Interactions And Ligand Screening For Protein Separation

Human serum albumin(HSA)is an extracellular protein with the highest concentration in blood plasma,which is also a carrier for many small molecules and has an exceptional binding capacity for many endogenous and exogenous ligands.HSA is excess-demand with high medical value.In this work Site?of HSA was focused,and the methods of molecular docking and molecular dynamics simulation were used to investigate binding mode and dynamic binding process between HSA and ligands.In addition,the compounds containing indole ring in the small molecule database were screened as the potential ligand for HSA with molecular docking.New resin with the selected ligand was prepared,and the performance of HSA adsorption was evaluated.The results would guide the design of new ligands for the efficient separation of HSA.Main results were summarized as follows.Firstly,the molecular interactions between Site?of HSA and twelve Site?-specific ligands were evaluated with molecular simulation methods.The results showed that hydrophobic interactions were the main driving force for binding and the electrostatic interactions played a secondary role.The key residues and binding mode on Site?were identified with the computational alanine-scanning approach.Three-layer binding mode was found from the entrance to the interior of the binding pocket,contributing electrostatic interactions,hydrophobic interactions and mixed interactions,respectively.Molecular docking and molecular dynamics simulation were also used to predict the binding mode of L-tryptophan on Site?.Secondly,molecular simulation methods were used to investigate the dynamic binding process of ibuprofen to Site?of HSA.The binding process could be divided into four phases:long-range attraction,adjustment on the surface,entering to Site?pocket;and stable binding at Site?.After evaluating van der Waals 'and electrostatic interaction energies during the binding process,it was found that the initial major driving force involved electrostatic attractions.Subsequently,ibuprofen was locked between two polar regions on the surface near Site?and then moved to Site?;Ibuprofen then entered the pocket of Site?by combinatorial effects of polar and hydrophobic residues nearby the entrance of Site?Finally,electrostatic and hydrophobic interactions formed the stable binding of ibuprofen in Site?.The molecular surface near Site?was observed to change significantly during binding,which indicates an induced-fit mechanism.Finally,the Site?-specific ligands were screened with molecular simulation methods.9556 small molecules containing indole ring were obtained from ZINC,a small molecule database,and docked onto Site?of HSA by DOCK software.After eliminating the influence of molecular weight,the results showed that the more electro-negative groups,the more donor and acceptor of hydrogen bonds,the less rings,the stronger binding to Site?of HSA was found.CDOCKER was used further to evaluate top-2000 small molecules from DOCK for the next screening step.Based on the analysis and comparison,N-acetyl-L-tryptophan(L-NAT)was selected as the appropriate ligand.New resin with L-NAT as the functional ligands and agarose gel as the matrix was prepared.A typical pH-dependent adsorption and high adsorption capacity was found.The highest saturated adsorption capacity Qm was 124.05 mg/g gel at pH 5,and the dissociation constant Kd was 0.015 mg/ml.The results showed the new resin had a good potential for HSA separation.In the work,molecular simulation methods were used to evaluate the binding mode on site?of HSA,analyze the dynamic process of ibuprofen binding and conduct the high throughput screening of Site?-specific ligands.The efficiency of the ligand selected was confirmed.The results demonstrated that molecular simulation methods could provide an appropriate guidance for the screening and design of the site-specific ligands.