Engineered zwitterionic phosphorylcholine monolayers for elucidating multivalent binding kinetics of C-reactive protein

What is it about?

Understanding of the activation dynamics of C-reactive protein (CRP) on plasma membranes is important in the development of zwitterionic biomaterials for their uses in the tissues of inflammation and infection. Previously, the use of a zwitterionic phosphorylcholine group, a biomimetic ligand for CRP in the presence of calcium ions, for binding experiments has revealed that the adsorption dynamics changed by ionic microenvironments. Here we focused on the effect of the ligand density on a surface, a major physicochemical parameter, on the multivalent binding modes. A building block from synthetic origin, a phospholipid analogue with thiol ends, was developed for making a cell membrane-mimicked self-assembled monolayers with tunable lateral ligand density on the molecular basis. The multivalent binding kinetics of CRP, a pentraxin in the original conformation, onto the engineered surface was measured using a surface plasmon resonance technique. The binding experiments revealed that the on-rate and off-rate constants in the first ligand-occupation reaction increased with increasing the ligand density, which resulted in stable values of the dissociation constant. Notably, the binding affinity in the second ligand-occupation reaction showed the optimal value as a function of the ligand density. Moreover, the binding experiments using a monomeric CRP-specific DNA aptamer revealed that pentameric CRP underwent structural transition into the monomers following the adsorption onto the surfaces via multivalent contacts in a pH-dependent manner. The bioengineering-based approach reveals for the first time how the multiple binding reaction is altered by the ligand arrangement at the molecular resolution and how CRP is activated by the conformational transition induced by the multiplex bindings.

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Why is it important?

C-reactive protein (CRP), a major acute-phase pentraxin, binds to plasma membranes through the multivalent contacts with zwitterionic phosphorylcholine groups. However, details in the molecular dynamics is unknown due to a lack of proper sensing platform. The paper describe the synthesis of thiol-functionalized phosphorylcholine for the development of a robust cell membrane-mimetic surface on a surface plasmon resonance sensor at desired lateral ligand densities. The engineered approach on molecular basis enables a rigorous arrangement of the ligand on the surface, whose tunability and robustness are not achieved using conventional supported lipid layers. The effect of the ligand density on the multivalent binding kinetics provides the understanding of how the multivalent contacts induce conformational transitions of CRP and responses to inflammation.