Metal-ion binding to parvalbumin. A 113Cd-n.m.r. study of the binding of different lanthanide ions.

113Cd-n.m.r. studies were used to investigate the binding of the lanthanide ions La3+, Gd3+, Tb3+, Yb3+ and Lu3+ to parvalbumins. It was shown that lanthanide ions with a smaller ionic radius bind sequentially to Cd2+-saturated parvalbumin, whereas those with a larger ionic radius bind with similar affinity to both the CD site and the EF site. The smallest ion, Lu3+, does in fact not compete significantly with Cd2+ for the CD site in carp parvalbumin, but appears to bind only to the EF site. This preference of the smaller lanthanide ions for the EF site was used to assign the n.m.r. signals for protein-bound 113Cd. By using Cd n.m.r. and Tb3+ fluorescence it was also shown for alpha-lineage parvalbumin from pike that these proteins possess a third site that can bind lanthanide ions. This site is, however, much weaker than in the beta-lineage parvalbumins. It was used to assign the 113Cd resonances from protein-bound Cd2+ ions in the spectrum of pike pI5.0 parvalbumin.

Observation of the 43Ca NMR signals from Ca2+ ions bound to calmodulin, parvalbumin, and troponin C.

Through the combined use of isotopically enriched 43Ca2+, high magnetic field, Fourier transform techniques, and a high-performance probe, the 43Ca NMR signals from ions bound to three proteins have been observed. These proteins are calmodulin, parvalbumin, and troponin C. Both longitudinal (1/T1) and transverse (1/T2) relaxation rates were measured for protein-bound 43Ca2+, and the quadrupole coupling constant and correlation time were obtained from these data. The calculated correlation times are in good agreement with the rotational correlation time for the entire protein molecules, which indicates that the Ca2+-binding sites are relatively rigid.

NMR studies on parvalbumin phylogeny and ionic interactions.

The inspection of several muscular parvalbumins from different species by two NMR methods (113Cd resonance and 1H relaxation measurements) allows two classes of parvalbumins to be distinguished according to their ion-binding properties. This result is in agreement with the phylogenetic classification of parvalbumins in two series, alpha and beta, which was established on the basis of the primary structures of these proteins. All parvalbumins are characterized by the presence of two primary cationic sites CD and EF, with structural features closely related to those already known on the basis of X-ray crystallographic studies of the beta parvalbumin pI 4.25 from carp muscle. However, parvalbumins of the beta series are characterized by a secondary cation (Ca2+, Mg2+ and other cations) binding site which is absent (or at least inaccessible) in parvalbumins of the alpha series. The major component from thornback ray (pI 4.45) behaves as an alpha parvalbumin as shown by the present NMR studies, although its primary structure suggests a closer similarity with the parvalbumins of the beta series.