Fig. 1.

UDCA binds a single site in IBABP. (A) Tryptophan fluorescence spectroscopy was used to quantify the binding affinity of UDCA to IBABP. IBABP (250 µl at 10 µM) was titrated in a stepwise manner with 1 µl increments of UDCA (5 mM). The specific binding measured by normalized changes in emission fluorescence is plotted against UDCA concentration. The hyperbolic binding curve indicates IBABP can bind UDCA in a single binding site. (B, C) The binding sites of UDCA and CA to IBABP were determined by ligand-observed ¹H, ¹⁵N HSQC NMR spectroscopy. The contour plots of NMR spectra of ¹⁵N-GCA (B) or ¹⁵N-GUDCA (C) bound to IBABP at a three-to-one molar ratio are shown. Arrows indicate peaks corresponding to bound bile acids.