Investigation of binding behavior of alpha- and beta-ionones to beta-lactoglobulin at different pH values using a diffusion-based NOE pumping technique.
Diffusion-based NMR techniques were employed to study effects of pH on beta-lactoglobulin (BLG) conformation and binding affinity to alpha- and beta-ionone. In the first part of the study, the influence of pH on the diffusion coefficient of BLG in D(2)O solution was investigated using a stimulated-echo NMR experiment. The diffusion coefficient of BLG decreased with increasing pH values. A significant decrease in the diffusion coefficient observed at pH 11 may be due to total unfolding (denaturation) of the protein, resulting in hydrophobically driven self-aggregation. A diffusion-based NOE pumping technique was then applied to determine the relative binding affinities between alpha- and beta-ionones and BLG at pH values varying from 3 to 11. An increase in signal intensities for beta-ionone with increasing molar concentration ratios between beta-ionone and BLG was observed at all pH ranges studied. The increased signal intensities reflect increased relative binding affinity. The greatest binding affinity occurred at pH 9 and the lowest at pH 11. alpha-Ionone showed binding evidence only at pH 9, and the binding was significantly weaker than that obtained for beta-ionone at the same pH. The high affinity observed for both aroma compounds at pH 9 may be due to a flexible conformation of BLG at this pH so that the flavor ligand accessibility increases. Conversely, alkaline denaturation occurring at pH 11 gives rise to relatively lower binding affinity compared to that observed at the other pH values.
Jung
Da-Mi
DM
Department of Viticulture and Enology, University of California at Davis, Davis, California 95616-8749, USA.
Ebeler
Susan E
SE
D004058:Diffusion; D006863:Hydrogen-Ion Concentration; D007782:Lactoglobulins / Q000737:chemistry* / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D045792:Norisoprenoids; D011487:Protein Conformation; D013729:Terpenes / Q000378:metabolism*
D007782:Lactoglobulins; D045792:Norisoprenoids; D013729:Terpenes; C008157:beta-ionone; C011879:alpha-ionone