Capillary electrophoretic study on pH dependence of enantioselective disopyramide binding to genetic variants of human alpha1-acid glycoprotein.
A high-performance frontal analysis-capillary electrophoresis (HPFA-CE) method was applied to investigate the effect of pH on the drug binding properties of genetic variants of human alpha1-acid glycoprotein (AGP), A variant and a mixture of F1S variants. The unbound concentrations of a model basic drug, disopyramide (DP), in A variant solutions and in F1S variant solutions were measured by HPFA-CE to evaluate binding constants at pH 4.0, 5.0, 6.0 and 7.4. The binding between DP and A variant was gradually weakened by acidification of background buffer (from pH 7.4 to 4.0), while the binding between DP and FIS variants decreased at first (from pH 7.4 to 6.0), and then gained (from pH 6.0 to 4.0). Consequently, DP was more strongly bound to A variant than to FIS variants at pH 7.4, while at pH 4.0 DP was more strongly bound to F1S variants. At any pH (S)-DP was bound more strongly than (R)-DP, and the enantioselectivity of A variant was significantly higher than that of F1S variants. Electrophoretic mobilities of the AGP genetic variants decreased along with a decrease in pH. Fluorescent emission of these genetic variants indicated a distinct conformational change between pH 5.0 and 4.0. However, there was no significant difference in the electrophoretic mobility and the fluorescent emission spectrum between these variants at any pH. On the other hand, circular dichroism analyses revealed that beta-sheet content in FIS variants diminished as pH decreased, while that in A variant increased. These results suggest that the conformational change induced by acidification of background buffer differs between these genetic variants, and this causes the difference in DP bindability.
Kuroda
Yukihiro
Y
Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan. kurodada@pharm.kyoto-u.ac.jp
Matsumoto
Shuhei
S
Shibukawa
Akimasa
A
Nakagawa
Terumichi
T
D000889:Anti-Arrhythmia Agents / Q000737:chemistry*; D004206:Disopyramide / Q000737:chemistry*; D019075:Electrophoresis, Capillary / Q000379:methods; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009961:Orosomucoid / Q000737:chemistry / Q000235:genetics*
D000889:Anti-Arrhythmia Agents; D009961:Orosomucoid; D004206:Disopyramide