Abstract
We have measured the binding of two radioiodinated monoclonal anti-dinitrophenyl antibodies (IgE and IgG2a) to two dinitrophenylated lipid haptens in lipid bilayer membranes having various compositions and physical properties. These antibodies bind strongly to the lipophilic dinitrophenyl group in some membranes. Dimyristoylphosphatidylcholine and dipentadecanoylphosphatidylcholine containing 2 mol % dinitrophenyl lipid hapten bind anti-dinitrophenyl antibodies below the chain-melting transition temperatures of these lipids (22 and 35 degrees C, respectively) but not above these temperatures. Evidently, the lipophilic dinitrophenyl group is partially or completely buried in the hydrophobic region of these bilayers at temperatures above the chain-melting transition temperatures. The inclusion of increasing concentrations of cholesterol in such membranes (e.g. in dimyristoylphosphatidylcholine at 37 degrees C) results in a marked enhancement of antibody binding. It was found that a third lipid hapten containing the dinitrophenyl group does not show this strong dependence of antibody binding on the physical state of the lipid membrane. The weak immunologic degranulation of rat basophil leukemia cells by dimyristoylphosphatidylcholine membrane targets at 37 degrees C can be attributed to a weak binding of anti-dinitrophenyl IgE to these membranes (Balakrishnan, K., Hsu, F. J., Cooper, A. D., and McConnell, H. M. (1982) J. Biol. Chem. 257, 6427-6433). However, if the antibody is first allowed to bind to this membrane below the lipid chain-melting transition temperature, these IgE-coated membrane targets are very effective in releasing serotonin from the rat basophil leukemia cells when the temperature is raised to 37 degrees C.
Highlights
IgG2,) to two dinitrophenylated lipid haptens in lipid % Dnp’ lipidhapten evoked onlya weak immunologicdegranbilayer membranes having various compositions and physical properties
Be-The weak immunologic degranulation of rat baso- cause specific antigen-antibody binding is an early step in all phil leukemia cells by dimyristoylphosphatidylcholine these immunological responses, an examination of anti-Dnp membrane targets at37 “Ccan be attributed to aweak antibody binding to Dnp lipid haptens in various lipid membinding of anti-dinitrophenyl IgE to these membranes branes was undertaken
We have not attempted aquantitative analysis of these kinetic data. This observation, that anti-Dnp IgE bound to liposomes remains associated with them for about 10-15 min, suggested that we might observe stimulation of RBL cells by DMPC membrane targets which had beenpreviously coated with anti-Dnp IgE below the transition temperature
Summary
Binding of Iodinated Antibodies toSolid and Fluid Membrane Targets-The binding of ['251]anti-DnpIgE to DMPC, DPPC, DSPC, and DBPC membrane targets containing mol % Dnp lipid haptenI was measuredat 37 "C.Antibody binding to the lipid haptens in the DMPC liposomes was weak and almostunmeasurable. On a Scatchard plot, all the three solid liposomes yieldedastraight line withaslope or an apparent affinity constant of 2 k 0.2 X 10' M" (Fig. 1). The binding of ['251]anti-Dnp IgE to DMPC liposomes at 0 "C (solid) was studied and yielded an apparent affmity constant of 7 f 1 X lo M-*, which was similar to that obtained with DPPC liposomes at 0 "C, uiz. Apparent affinity constants KI of [125Z/anti-nitroxidZegGl to the nitroxide lipid haptenZI in fluid andsolid membrane targets
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