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 surface properties. Substantial differences in the electrostatic forces measured with denatured Fab' further demonstrated that the measured electrostatic surface properties and the consequent long-range interaction forces are controlled by the protein surface composition. At short range, the strength of the Fab'-mediated adhesion was modulated not only by the length of the fluorescein tether but also by membrane hydration. Steric hydration barriers at the membrane surface reduced the adhesion strength in proportion to their range of influence. These results provide direct evidence that long-range protein interactions with immobilized ligands are controlled by both the protein and the membrane surface compositions, while short-range, specific binding is modulated by both the protein structure and the membrane interfacial properties.
Leckband
D E
DE
Department of Chemical Engineering, University of Illinois at Urbana, Champaign 61801, USA.
Kuhl
T
T
Wang
H K
HK
Herron
J
J
Müller
W
W
Ringsdorf
H
H
D000911:Antibodies, Monoclonal / Q000276:immunology*; D000937:Antigen-Antibody Reactions; D002462:Cell Membrane / Q000276:immunology; D004563:Electrochemistry; D019793:Fluorescein; D005452:Fluoresceins; D006241:Haptens / Q000276:immunology*; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D007074:Immunoglobulin G / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D011487:Protein Conformation; D011489:Protein Denaturation; D013499:Surface Properties
D000911:Antibodies, Monoclonal; D005452:Fluoresceins; D006241:Haptens; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D019793:Fluorescein