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Identification of the ferredoxin interaction sites on ferredoxin-dependent glutamate synthase from Synechocystis sp. PCC 6803

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Abstract

Based on in silico docking methods, five amino acids in glutamate synthase (Gln-467, His-1144, Asn-1147, Arg-1162, and Trp-676) likely constitute key binding residues in the interface of a glutamate synthase:ferredoxin complex. Although all interfacial mutants studied showed the ability to form a complex under low ionic strength, these docking mutations showed significantly less ferredoxin-dependent activities, while still retaining enzymatic activity. Furthermore, isothermal titration calorimetry showed a possible 1:2 molar ratio between the wild-type glutamate synthase and ferredoxin. However, each of our interfacial mutants showed only a 1:1 complex with ferredoxin, suggesting that the mutations directly affect the glutamate synthase:ferredoxin heterodimer interface.

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Acknowledgements

We especially thank Prof. Guy Hanke (Queen Mary University of London) for reading this manuscript and suggesting layout of this manuscript. This research was supported by AR063634 (to R.B.S) and the Office of Basic Energy Sciences of the U.S. Department of Energy, through Grant DE-FG03-99ER20346 (to D.B.K.).

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Author notes

  1. Jacaranda Solis

    Present address: Immunology and Molecular Microbiology, Texas Tech University Health Science Center, Lubbock, TX, 79430-6591, USA

  2. Nanditha Vaidyanathan

    Present address: Depuy Synthes Companies, 1302 Wrights Lane East, West Chester, PA, 19380, USA

  3. Anurag P. Srivastava

    Present address: Department of Biochemistry and Molecular Biology, Rosalind Franklin University of Medicine and Science, North Chicago, Illinoi, 60064, USA

  4. David B. Knaff—Posthumous.

Authors and Affiliations

  1. Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, 79409-1061, USA

    Masakazu Hirasawa, Anurag P. Srivastava & David B. Knaff

  2. Center for Biotechnology and Genomics, Texas Tech University, Lubbock, TX, 79409-3132, USA

    Jacaranda Solis, Nanditha Vaidyanathan & David B. Knaff

  3. Departments of Internal Medicine and Biochemistry, University of Texas Southwestern Medical Center, Dallas, TX, 75390-9038, USA

    R. Max Wynn

  4. Department of Cell Physiology and Molecular Biophysics, Texas Tech Health Science Center, Lubbock, TX, 79430-6551, USA

    Roger B. Sutton

Authors
  1. Masakazu Hirasawa
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Contributions

MH and DBK conceived of and designed the studies. JS and NV provided all variants, AS provided ferredoxin, RBS contributed in silico docking model, and MH and RMW analyzed the data and wrote the manuscript with input from all authors.

Corresponding author

Correspondence to R. Max Wynn.

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Hirasawa, M., Solis, J., Vaidyanathan, N. et al. Identification of the ferredoxin interaction sites on ferredoxin-dependent glutamate synthase from Synechocystis sp. PCC 6803. Photosynth Res 134, 317–328 (2017). https://doi.org/10.1007/s11120-017-0446-z

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  • DOI: https://doi.org/10.1007/s11120-017-0446-z

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