Binding Site Classification
The example shown in Fig. 3.9 demonstrates this technique for the HSA ligand warfarin in competition with its stable isotope-labeled congener warfarin-D6. Here, the concentration of the receptor and deuterated ligand are held constant while warfarin is added at increasing concentrations. The ALIS MS response of warfarin increases while the response of warfarin-D6 diminishes as it is competed from its binding site on HSA. This is an absolute example of direct binding competition (Scheme 3.1), since the labeled and unlabeled compounds bind the same site with identical affinities, yet are distinguishable by their different molecular weights. Importantly, if two compounds bind the same site, the ratio of the ALIS
05 Intensity iog| Warfarin I, ijM
Fig. 3.9 ALIS-MS results for the titration of 5 mM HSA with warfarin in the presence of a 5 mm concentration of its stable isotope-labeled congener warfarin-D6. Increasing concentrations of warfarin reduce the response of warfarin-D6 due to isosteric binding competition. Reprinted from  with permission from the American Chemical Society.
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