The effects of anions on the solution structure of Na,K-ATPase.
Anions interact with protein to induce structural changes at ligand binding sites. The effects of anion complexation include structural stabilization and promote cation-protein interaction. This study was designed to examine the interaction of aspirin and ascorbate anions with the Na+, K+-dependent adenosine triphosphatase (Na,K-ATPase) in H2O and D2O solutions at physiological pH, using anion concentrations of 0.1 microM to 1 mM with final protein concentration of 0.5 to 1 mg/ml. Absorption spectra and Fourier transform infrared (FTIR) difference spectroscopy with its self-deconvolution, second derivative resolution enhancement and curve-fitting procedures were applied to characterize the anion binding mode, binding constant, and the protein secondary structure in the anion-ATPase complexes. Spectroscopic evidence showed that the anion interaction is mainly through the polypeptide C=O and C-N groups with minor perturbation of the lipid moiety. Evidence for this came from major spectral changes (intensity variations) of the protein amide I and amide II vibrations at 1651 and 1550 cm(-1). respectively. The anion-ATPase binding constants were K=6.45 x 10(3) M(-1) for aspirin and K=1.04 x 10(4) M(-1) for ascorbate complexes. The anion interaction resulted in major protein secondary structural changes from that of the alpha-helix 19.8%; beta-pleated sheet 25.6%; turn 9.1%; beta-antiparallel 7.5% and random 38% in the free Na,K-ATPase to that of the alpha-helix 24-26%; beta-pleated 17-18%; turn 8%; beta-antiparallel 5-3% and random 45.0% in the anion-ATPase complexes.
Neault
J F
JF
Department of Chemistry-Biology, University of Québec at Trois-Rivières, Canada.
Benkiran
A
A
Malonga
H
H
Tajmir-Riahi
H A
HA
D000818:Animals; D000838:Anions; D001205:Ascorbic Acid / Q000737:chemistry; D001241:Aspirin / Q000737:chemistry; D001665:Binding Sites; D006168:Guinea Pigs; D066298:In Vitro Techniques; D046911:Macromolecular Substances; D017433:Protein Structure, Secondary; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry*; D012996:Solutions; D013053:Spectrophotometry; D017550:Spectroscopy, Fourier Transform Infrared
D000838:Anions; D046911:Macromolecular Substances; D012996:Solutions; D000254:Sodium-Potassium-Exchanging ATPase; D001205:Ascorbic Acid; D001241:Aspirin