Conformational Change of H+-ATPase Monomer Revealed on Segmental Isotope Labeling NMR Spectroscopy
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- 作者：Hiromasa Yagi ; Takuya Tsujimoto ; Toshio Yamazaki ; Masasuke Yoshida ; and Hideo Akutsu
- 刊名：Journal of the American Chemical Society
- 出版年：2004
- 出版时间：December 22, 2004
- 年：2004
- 卷：126
- 期：50
- 页码：16632 - 16638
- 全文大小：343K
- 年卷期：v.126,no.50(December 22, 2004)
- ISSN：1520-5126
文摘
F1-ATPase has been shown to be a stepwise molecular motor. Its rotation mechanism has beenexplained by the interaction of the 
axis with the open and closed forms of the 
subunit. Although NMRshould be a powerful method for elucidating its mechanism, its molecular size (473 amino acid residues,52 kDa) is a major obstacle. We have applied segmental labeling based on intein ligation to the subunit,and succeeded in assigning 89% of the NH (402/451), 89% of the C
(417/473), 83% of the C (357/431),and 90% of the CO (425/473) signals of the subunit monomer. The secondary structures predicted fromthe chemical shifts of the main chain atoms and the relative orientations determined from residual dipolarcouplings indicated that the subunit monomer takes on the open form in the absence of nucleotide.Furthermore, the chemical shift perturbation and the residual-dipolar-coupling changes induced by nucleotidebinding show that conformational change from the open to the closed form takes place on nucleotide binding.The intrinsic conformational change of the subunit monomer induced by nucleotide binding must be oneof the essential driving forces for the rotation of F1-ATPase.
axis with the open and closed forms of the subunit. Although NMRshould be a powerful method for elucidating its mechanism, its molecular size (473 amino acid residues,52 kDa) is a major obstacle. We have applied segmental labeling based on intein ligation to the subunit,and succeeded in assigning 89% of the NH (402/451), 89% of the C (417/473), 83% of the C (357/431),and 90% of the CO (425/473) signals of the subunit monomer. The secondary structures predicted fromthe chemical shifts of the main chain atoms and the relative orientations determined from residual dipolarcouplings indicated that the subunit monomer takes on the open form in the absence of nucleotide.Furthermore, the chemical shift perturbation and the residual-dipolar-coupling changes induced by nucleotidebinding show that conformational change from the open to the closed form takes place on nucleotide binding.The intrinsic conformational change of the subunit monomer induced by nucleotide binding must be oneof the essential driving forces for the rotation of F1-ATPase.