Differential binding of disopyramide and warfarin enantiomers to human α1-acid glycoprotein variants

Authors


K. Miyazaki PhD, Department of Pharmacy, Hokkaido University Hospital, School of Medicine, Hokkaido University, Kita 14-jo, Nishi 5-chome, Kita-ku, Sapporo 060–8648, Japan. Tel.: + 81 11 706 5680; Fax: + 81 11 756 1505; E-mail: katsumim@med.hokudai.ac.jp

Abstract

Aims  The F1S and A genetic variants of α1-acid glycoprotein (AAG) change under various physiological and pathological conditions. They also vary in their drug binding abilities. We have studied the stereoselective binding ability of each of the AAG variants using enantiomers of disopyramide (DP) and warfarin (WR).

Methods  The AAG variants were separated by hydroxyapatite chromatography. Binding of drug enantiomers to the AAG variants was studied by the Hummel–Dreyer method. The characteristics of the binding activities were examined by Scatchard plot analysis. The first five amino-terminal amino acids (residues 112–116) of the cyanogen bromide (CNBr) fragment (residues 112–181) of each of the separated AAG fractions were elucidated by Edman degradation.

Results  Commercial AAG was separated into two main fractions. Residues 112–116 of fraction 2 were identical to the amino acid sequences predicted from the AAG A gene, LAFDV, and encode the F1S variant. In fraction 3, the deduced amino acid sequence of the AAG B gene, FGSYL, was established, and encodes the A variant. The binding affinities of both DP enantiomers in fraction 3 were significantly higher than those in fraction 2. The differences between dissociation constants (Kd) in fractions 2 and 3 were 5.2-fold for (S)-DP (P < 0.05) and 3.7-fold for (R)-DP (P < 0.001). The dissociation constant of (S)-DP (0.39 ± 0.08 µm) was lower than that of (R)-DP (0.53 ± 0.10 µm) in fraction 3 [95% confidence interval (CI) − 0.282, − 0.010; P < 0.05], although the binding activities of the DP enantiomers were almost the same in fraction 2. By contrast WR enantiomers had a higher binding affinity in fraction 2 than in fraction 3, the differences in dissociation constants between fractions 2 and 3 being 12.6-fold for (S)-WR (P < 0.001) and 8.3-fold for (R)-WR (P < 0.001). The dissociation constant of (S)-WR (0.28 ± 0.10 µm) was significantly lower than that of (R)-WR (0.48 ± 0.08 µm) in fraction 2 (95% CI − 0.369, − 0.028; P < 0.05), but there were no significant differences between the binding activities of WR enantiomers in fraction 3.

Conclusions  DP and WR enantiomers bind preferentially to fraction 3 and fraction 2, respectively. Fractions 2 and 3 are encoded by the AAG A and the AAG B genes, respectively.

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