Figure 5
Binding affinity and identification of contact residues of nIgG peptides with ficolin.
Surface plasmon resonance (response unit) to characterize the binding affinities between nIgG peptides and H-ficolin, under normal and infection-inflammation conditions. GlcNAc-BSA was immobilized on the CM5 chip to expose GlcNAc as a ligand for anchoring ficolin to the chip. H-ficolin (200β nM) was first injected over GlcNAc, respectively for 100β s (association time) followed by buffer flow (wash) for 200β s (dissociation time). Increasing concentrations of nIgG peptides (5, 10, 20 and 50β ΞΌM) were injected over the H-ficolin bound to the chip, under similar conditions. Mutant peptides (with Arg, Lys or His substituted to Ala) did not bind to the corresponding proteins. Human serum albumin (HSA) injected after H-Ficolin served as a negative control (blue, see top 2 panels). nIgG peptides (WT or mutants) are tabulated below the respective graphs. Peptides, injected directly over the GlcNAc-immobilized chip as controls, showed no binding to GlcNAc (blue). Data were analyzed using BIAevaluation software 3.2. The binding curves (black) are overlaid with the fit of 1:1 interaction model (red). The plots are a typical representation of 3 independent experiments.
