Homareda H, Ishii T, Takeyasu K
First Department of Biochemistry, Kyorin University School of Medicine, Tokyo 181-8611, Mitaka, Japan.
Eur J Pharmacol. 2000 Jul 21;400(2-3):177-83. doi: 10.1016/s0014-2999(00)00411-8.
Oligomycin inhibits Na(+),K(+)-ATPase activity by stabilizing the Na(+) occlusion but not the K(+) occlusion. To locate the binding domain of oligomycin on Na(+),K(+)-ATPase, the tryptic-digestion profile of Na(+),K(+)-ATPase was compared with the profile of Na(+) occlusion within the digested Na(+),K(+)-ATPase in the presence of oligomycin. The Na(+) occlusion profile is responsible for the digestion profile of the alpha-subunit, which is the catalytic subunit of the ATPase. The effect of oligomycin on chimeric Ca(2+)-ATPase activity was examined. The chimera used, in which the 163 N-terminal amino acids of chicken sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase 1 were replaced with the 200 N-terminal amino acids of the chicken Na(+),K(+)-ATPase alpha1-subunit, partially retains the Na(+)-dependent characteristics of Na(+), K(+)-ATPase, because the chimeric Ca(2+)-ATPase activity is activated by Na(+) but inhibited by ouabain, a specific inhibitor of Na(+),K(+)-ATPase (Ishii, T., Lemas, M.V., Takeyasu, K., 1994, Proc. Natl. Acad. Sci. U. S. A., 91, 6103-6107). Oligomycin depressed the activation by Na(+) of the chimeric Ca(2+)-ATPase activity. These findings suggest that the 200 N-terminal amino acids of the Na(+), K(+)-ATPase alpha-subunit include a binding domain for oligomycin.
寡霉素通过稳定钠（Na⁺）封闭状态而非钾（K⁺）封闭状态来抑制钠钾（Na⁺,K⁺）-ATP酶活性。为了确定寡霉素在钠钾（Na⁺,K⁺）-ATP酶上的结合结构域，将钠钾（Na⁺,K⁺）-ATP酶的胰蛋白酶消化图谱与在寡霉素存在下消化后的钠钾（Na⁺,K⁺）-ATP酶中的钠（Na⁺）封闭图谱进行了比较。钠（Na⁺）封闭图谱决定了α亚基的消化图谱，α亚基是ATP酶的催化亚基。研究了寡霉素对嵌合型钙（Ca²⁺）-ATP酶活性的影响。所使用的嵌合体中，鸡肌浆网/内质网钙（Ca²⁺）-ATP酶1的163个N端氨基酸被鸡钠钾（Na⁺,K⁺）-ATP酶α1亚基的200个N端氨基酸所取代，该嵌合体部分保留了钠钾（Na⁺,K⁺）-ATP酶的钠（Na⁺）依赖性特征，因为嵌合型钙（Ca²⁺）-ATP酶活性被钠（Na⁺）激活但被哇巴因抑制，哇巴因是钠钾（Na⁺,K⁺）-ATP酶的特异性抑制剂（石井，T.，莱马斯-M.V.，竹安，K.，1994年，美国国家科学院院刊，91，6103 - 6107）。寡霉素降低了钠（Na⁺）对嵌合型钙（Ca²⁺）-ATP酶活性的激活作用。这些发现表明，钠钾（Na⁺,K⁺）-ATP酶α亚基的200个N端氨基酸包含寡霉素的结合结构域。
