0000000000165148
AUTHOR
James N. Herron
4-4-20 anti-fluorescyl IgG Fab' recognition of membrane bound hapten: direct evidence for the role of protein and interfacial structure.
The surface forces apparatus was used to identify the molecular forces that control the interactions of monoclonal 4-4-20 antifluorescyl IgG Fab' fragments with fluorescein-presenting supported planar bilayers. At long range, the electrostatic force between oriented Fab' and fluorescein monolayers was controlled by the composition of the protein exterior surrounding the antigen-combining site rather than by the overall protein charge. The measured positive electrostatic potential of the Fab' monolayer at pH > pI(Fab') was consistent with the structure of the exposed Fab' surface in which a ring of positive charge at the mouth of the antigen-combining site dominates the local electrostatic s…
Binding, Interaction, and Organization of Proteins with Lipid Model Membranes
Model membrane systems are used to investigate protein recognition and binding at interfaces. Fluorescence microscopy results are presented for interactions of the proteins, phospholipase A2 and antifluorescyl IgG, at lipid monolayer interfaces. Total internal reflection fluorescence measurements are used to quantify albumin and IgG adsorption to supported lipid monolayers.
Force probe measurements of antibody-antigen interactions.
The surface force apparatus has been used to quantify directly the forces that govern the interactions between proteins and ligands. In this work, we describe the measured interactions between the antigen fluorescein and the Fab' fragment of the monoclonal 4-4-20 anti-fluorescyl IgG antibody. Here we first describe the use of the surface force apparatus to demonstrate directly the impact of the charge composition in the region of the antibody binding site on the antibody interactions. Several approaches are described for immobilizing antigens, antibodies, and proteins in general for direct force measurements. The measured force profiles presented are accompanied by an extensive discussion o…
Quenching of fluorescein-conjugated lipids by antibodies. Quantitative recognition and binding of lipid-bound haptens in biomembrane models, formation of two-dimensional protein domains and molecular dynamics simulations
Three model biomembrane systems, monolayers, micelles, and vesicles, have been used to study the influence of chemical and physical variables of hapten presentation at membrane interfaces on antibody binding. Hapten recognition and binding were monitored for the anti-fluorescein monoclonal antibody 4–4-20 generated against the hapten, fluorescein, in these membrane models as a function of fluorescein-conjugated lipid architecture. Specific recognition and binding in this system are conveniently monitored by quenching of fluorescein emission upon penetration of fluorescein into the antibody's active site. Lipid structure was shown to play a large role in affecting antibody quenching. Interes…