113Cd nuclear magnetic resonance study of the binding of Cd 2+ and Lu 3+ to silver hake parvalbumin

Abstract

The binding of Cd 2+ and Lu 3+ to a parvalbumin isotype isolated from silver hake (Merluccius bilinearis), SHPV-B, has been studied by 113Cd nuclear magnetic resonance (NMR). Titration at pH 6.7, with 113CdCl 2, of either apo SHPV-B or native SHPV-B ( i.e. Ca 2+ loaded) resulted in the emergence of two 113Cd resonances in the 113Cd NMR spectrum, one at about −94 ppm and another at about −97 ppm (relative to a 113 Cd 2+ signal at 0 ppm arising from (unenriched) 2 M CdSO 4, consistent with the presence of two distinct binding site environments on the protein for cadmium ion. Addition of Lu 3+ to 113Cd 2+ -substituted SHPV-B resulted in the preferential loss of the low field ( i.e. −94 ppm) 113Cd NMR signal, consistent with the existence of one highly selective site on the silver hake parvalbumin (for the binding of ions that are very close to Ca 2+ in their ionic radii and charge density, such as Cd 2+ and a second, less selective site that can accommodate ions of slightly differing size (and charge density). Comparison of our Lu 3+ addition results (on the silver hake parvalbumin) with the results of similar studies in the literature on pike and carp parvalbumin indicates significant similarities with the former and significant differences from the latter. These studies may bear on the evolutionary relatedness of parvalbumins from these three species.