Purine but not Pyrimidine Nucleotides Support Rotation of F 1-ATPase

Hiroyuki Noji, Dirk Bald, Ryohei Yasuda, Hiroyasu Itoh, Masasuke Yoshida, Kazuhiko Kinosita

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Abstract

The binding change model for the F1,-ATPase predicts that its rotation is intimately correlated with the changes in the affinities of the three catalytic sites for nucleotides. If so, subtle differences in the nucleotide structure may have pronounced effects on rotation. Here we show by single-molecule imaging that purine nucleotides ATP, GTP, and ITP support rotation but pyrimidine nucleotides UTP and CTP do not, suggesting that the extra ring in purine is indispensable for proper operation of this molecular motor. Although the three purine nucleotides were bound to the enzyme at different rates, all showed similar rotational characteristics: counterclockwise rotation, 120° steps each driven by hydrolysis of one nucleotide molecule, occasional back steps, rotary torque of ∼40 piconewtons (pN)·nm, and mechanical work done in a step of ∼80 pN·nm. These latter characteristics are likely to be determined by the rotational mechanism built in the protein structure, which purine nucleotides can energize. With ATP and GTP, rotation was observed even when the free energy of hydrolysis was-80 pN·nm/molecule, indicating -100% efficiency. Reconstituted FoF1-ATPase actively translocated protons by hydrolyzing ATP, GTP, and ITP, but CTP and UTP were not even hydrolyzed. Isolated F1 very slowly hydrolyzed UTP (but not CTP), suggesting possible uncoupling from rotation.

Original languageEnglish
Pages (from-to)25480-25486
Number of pages7
JournalJournal of Biological Chemistry
Volume276
Issue number27
DOIs
Publication statusPublished - 2001 Jul 6
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry

Cite this

Noji, H., Bald, D., Yasuda, R., Itoh, H., Yoshida, M., & Kinosita, K. (2001). Purine but not Pyrimidine Nucleotides Support Rotation of F 1-ATPase. Journal of Biological Chemistry, 276(27), 25480-25486. https://doi.org/10.1074/jbc.M102200200