Synthetic mimics of small Mammalian cell surface receptors

Receptors on the surface of mammalian cells promote the uptake of cell-impermeable ligands by receptor-mediated endocytosis. To mimic this process, we synthesized small molecules designed to project anti-dinitrophenyl antibody-binding motifs from the surface of living Jurkat lymphocytes. These synthetic receptors comprise N-alkyl derivatives of 3beta-cholesterylamine as the plasma membrane anchor linked to 2,4-dinitrophenyl (DNP) and structurally similar fluorescent 7-nitrobenz-2-oxa-1,3-diazole (NBD) headgroups. Insertion of two beta-alanine subunits between a DNP derivative and 3beta-cholesterylamine yielded a receptor that avidly associates with cell surfaces (cellular t(1/2) approximately 20 h). When added to Jurkat cells at 10 microM, this receptor enhanced uptake of an anti-DNP IgG ligand by approximately 200-fold in magnitude and approximately 400-fold in rate within 4 h (ligand internalization t(1/2) approximately 95 min at 37 degrees C). This non-natural receptor mimics many natural receptors by dynamically cycling between plasma membranes and intracellular endosomes (recycling t(1/2) approximately 3 min), targeting of protein ligands to proposed cholesterol and sphingolipid-enriched lipid raft membrane microdomains, and delivery of protein ligands to late endosomes/lysosomes. Quantitative dithionite quenching of fluorescent extracellular NBD headgroups demonstrated that other 3beta-cholesterylamine derivatives bearing fewer beta-alanines in the linker region or N-acyl derivatives of 3beta-cholesterylamine were less effective receptors due to more extensive trafficking to internal membranes. Synthetic cell surface receptors have potential applications as cellular probes, tools for drug delivery, and methods to deplete therapeutically important extracellular ligands.