1H-NMR studies of synthetic peptide analogues of calcium-binding site III of rabbit skeletal troponin C: effect on the lanthanum affinity of the interchange of aspartic acid and asparagine residues at the metal ion coordinating positions.
The present work determines the contribution of liganding aspartic acid (Asp) residues, at the +X, +Y, and +Z metal ion coordinating positions, to the lanthanum(3+) (La3+) ion binding affinity of synthetic analogues of calcium-binding site III of rabbit skeletal troponin C. Eight 13-residue synthetic analogues were prepared by solid-phase synthesis; the primary sequences of these analogues represent all possible combinations having aspartic acid and asparagine at the +X, +Y, and +Z positions. High-field proton nuclear magnetic resonance (NMR) spectroscopy was used to monitor the binding of the La3+ ion to each of the analogues. Comparison of the chemical shift changes showed large variations in the magnitude of the shift; these were reflected in the La3+ ion association constants determined for each analogue. The association constants ranged from 9.1 x 10(2) M-1 to 2.5 x 10(5) M-1. It was observed that those analogues with the larger number of acidic residues to coordinate the La3+ ion yielded the higher association constants. The La3+ ion binding results demonstrate that the Asp residues at the positions of study contribute equally and in an additive manner to the association constant and that the presence of neighboring Asp residues at either the +X and +Y, the +Y and +Z, or the +X and +Y and +Z metal ion coordinating positions introduced dentate-dentate repulsion, which, acts as to detract from the La3+ ion association constant of the analogues.
Department of Biochemistry, University of Alberta, Edmonton, Canada.
B J
BJ
Marsden
R S
RS
Hodges
B D
BD
Sykes
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001216:Asparagine / Q000378:metabolism*; D001224:Aspartic Acid / Q000378:metabolism*; D001665:Binding Sites; D002135:Calcium-Binding Proteins / Q000032:analysis*; D007700:Kinetics; D007811:Lanthanum / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D010446:Peptide Fragments / Q000138:chemical synthesis; D011817:Rabbits; D013816:Thermodynamics; D014336:Troponin / Q000032:analysis*; D019209:Troponin C
D000596:Amino Acids; D002135:Calcium-Binding Proteins; D010446:Peptide Fragments; D014336:Troponin; D019209:Troponin C; D001224:Aspartic Acid; D007811:Lanthanum; D001216:Asparagine