Bald D, Noji H, Yoshida M, Hirono-Hara Y, Hisabori T
PRESTO, Chemical Resources Laboratory, Tokyo Institute of Technology, Nagatsuta 4259, Japan.
J Biol Chem. 2001 Oct 26;276(43):39505-7. doi: 10.1074/jbc.C100436200. Epub 2001 Aug 22.
In F(1)-ATPase, the smallest known motor enzyme, unidirectional rotation of the central axis subunit gamma is coupled to ATP hydrolysis. In the present study, we report the redox switching of the rotation of this enzyme. For this purpose, the switch region from the gamma subunit of the redox-sensitive chloroplast F(1)-ATPase was introduced into the bacterial F(1)-ATPase. The ATPase activity of the obtained complex was increased up to 3-fold upon reduction (Bald, D., Noji, H., Stumpp, M. T., Yoshida, M. & Hisabori, T. (2000) J. Biol. Chem. 275, 12757-12762). Here, we successfully observed the modulation of rotation of gamma in this chimeric complex by changes in the redox conditions. In addition we revealed that the suppressed enzymatic activity of the oxidized F(1)-ATPase complex was characterized by more frequent long pauses in the rotation of the gamma subunit. These findings obtained by the single molecule analysis therefore provide new insights into the mechanisms of enzyme regulation.
在已知最小的动力酶F(1)-ATP酶中，中心轴亚基γ的单向旋转与ATP水解相偶联。在本研究中，我们报道了该酶旋转的氧化还原开关作用。为此，将氧化还原敏感的叶绿体F(1)-ATP酶γ亚基的开关区域引入细菌F(1)-ATP酶中。还原后，所得复合物的ATP酶活性提高了3倍（鲍德，D.，野地，H.，斯顿普，M.T.，吉田，M.和久座守，T.（2000年）《生物化学杂志》275，12757 - 12762）。在此，我们成功观察到通过氧化还原条件的变化对该嵌合复合物中γ旋转的调节作用。此外，我们还揭示了氧化的F(1)-ATP酶复合物被抑制的酶活性的特征是γ亚基旋转中更频繁出现长时间停顿。因此，通过单分子分析获得的这些发现为酶调节机制提供了新的见解。
