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| - Direct Observation of Ligand Binding to Membrane Proteinsin Living Cells by a Saturation Transfer Double Difference(STDD) NMR Spectroscopy Method Shows a SignificantlyHigher Affinity of Integrin αIIbβ3 in Native Plateletsthan in Liposomes
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| - About 30% of the proteins in mammalian systems are membrane bound or integrated (e.g.,GPCRs). It is inherently difficult to investigate receptor−ligand interactions on a molecular level in theirnatural membrane environment. Here, we present a new method based on saturation transfer difference(STD) NMR to characterize at an atomic level binding interactions of cell surface proteins in living cells.Implemented as a double difference technique, STD NMR allows the direct observation of binding eventsand the definition of the binding epitopes of ligands. The binding of the pentapeptide cyclo(RGDfV) to thesurface glycoprotein integrin αIIbβ3 of intact human blood platelets can be detected by saturation transferdouble difference (STDD) NMR in less than an hour. A 5-fold higher STD response reflects a significantlyhigher affinity of integrin αIIbβ3 in native platelets than in liposomes, which demonstrates the importance ofstudying membrane proteins in their natural environment. Also, the binding mode of cyclo(RGDfV) in thearginine glycine region is slightly different when interacting with native integrin in platelets compared tointegrin reintegrated into liposomes.
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