Adsorption of protein onto dye-ligand affinity adsorbent: Molecular dynamics simulation and experimental study

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Affinity chromatography is a well-established method for the identification, purification, and separation of macromolecules based on highly specific molecular recognition. Dye ligand affinity chromatography has been widely utilized for protein purification. Immobilized textile triazine dyes, particularly Cibacron blue 3GA (CB) has been used as affinity chromatography tools for protein purification for a long time. CB is able to bind various proteins. However, its interaction with a large number of seemingly unrelated proteins inevitably compromises its protein binding specificity and endows these molecules with a serious drawback. For an improved application of textile dyes in the separation of proteins, it is very important to understand the binding and specific mechanisms between dye ligand and proteins. In this project, serum albumin and hemoglobin which have been investigated in our previous works will be further studied as model proteins to investigate their adsorption onto dye ligand (CB) affinity chromatographic media. Due to the lack of experimental method that can detect the dynamic adsorption process within adsorbent pores, molecular dynamics (MD) simulations with both coarse grained and all atom models will be used for the first time in this study to investigate the dye ligand affinity chromatographic adsorption of proteins on a molecular scale. On the basis of the results of MD simulations and the former experimental research, both single and binary adsorption experiments of these proteins onto dye ligand affinity adsorbent will be performed under conditions which are rationally designed according to the binding mechanism learned from the MD simulations. A combination of MD simulations and adsorption experiments will give a comprehensive understanding of the binding and specific mechanisms of the affinity adsorption from both microscopic and macroscopic point of view. This understanding is a profound basis for the design and screening of new affinity dye ligands, as well as the design, optimization and control of the dye ligand affinity chromatography of proteins.