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The region from phenylalanine-28 to lysine-50 of a yeast mitochondrial ATPase inhibitor (IF1) forms an α-helix in solution

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Abstract

A mitochondrial ATPase inhibitor, IF1, is a 63 amino acid residue protein that regulates the activity of ATP synthase (F1Fo-ATPase). In the present study, we constructed mutant IF1 proteins with proline residues inserted into a wide range of their primary structures to determine the location and function of α-helix in the protein. A total of 11 yeast IF1 protein mutants were expressed and purified. Proline insertions in the region 28–50 reduced α-helical contents, indicating that the region formed a helix in solution. Oligomer formation of proline mutants at the C-terminal 38–60 region was markedly reduced, indicating that the region is required for oligomerization of the protein. Proline mutants at the N-terminal 18–39 region did not inhibit F1Fo-ATPase, indicating that the region is required for ATPase inhibitory activity. Inhibition of a proline insertion mutant between residues 44 and 45 that lost a large portion of the α-helix was slower, although the maximal inhibition level of the mutant protein was comparable to that of wild-type IF1. The results suggest that the helix of yeast IF1 facilitates binding to F1 by promoting initial interaction of the proteins.

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Acknowledgments

We thank Tomomi Maekawa, a technical staff member of Osaka City University, for performing DNA sequencing analysis. We thank Tomomi Shimonaka, a technical staff member of Osaka City University, for performing MALDI-TOF/MS analyses of IF1 proteins.

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Authors and Affiliations

  1. Department of Food and Human Health Sciences, Graduate School of Human Life Science, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi, Osaka, 558-8585, Japan

    Li Sun, Naomi Nakamae & Naoki Ichikawa

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  1. Li Sun
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  2. Naomi Nakamae
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  3. Naoki Ichikawa
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Correspondence to Naoki Ichikawa.

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Sun, L., Nakamae, N. & Ichikawa, N. The region from phenylalanine-28 to lysine-50 of a yeast mitochondrial ATPase inhibitor (IF1) forms an α-helix in solution. J Bioenerg Biomembr 47, 457–465 (2015). https://doi.org/10.1007/s10863-015-9627-7

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  • DOI: https://doi.org/10.1007/s10863-015-9627-7

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