Preprints
https://doi.org/10.5194/mr-2021-2
https://doi.org/10.5194/mr-2021-2

  20 Jan 2021

20 Jan 2021

Review status: this preprint is currently under review for the journal MR.

The long-standing relationship between Paramagnetic NMR and Iron-Sulfur proteins: the mitoNEET example. An old method for new stories or the other way around?

Francesca Camponeschi1,, Angelo Gallo2,, Mario Piccioli1,3, and Lucia Banci1,3 Francesca Camponeschi et al.
  • 1Consorzio Interuniversitario Risonanze Magnetiche MetalloProteine, Sesto Fiorentino, I-50019, Italy
  • 2Department of Pharmacy, University of Patras, Patras, GR-26504, Greece
  • 3Magnetic Resonance Center and Department of Chemistry, University of Florence, Sesto Fiorentino, I-50019, Italy
  • These authors contributed equally

Abstract. Paramagnetic NMR spectroscopy and iron-sulfur (Fe–S) proteins have maintained a synergic relationship for decades. Indeed, the hyperfine shifts with their temperature dependencies and the relaxation rates of nuclei of cluster-bound residues have been extensively used as a fingerprint of the type and of the oxidation state of the Fe–S cluster within the protein frame. The identification of NMR signals from residues surrounding the metal cofactor is crucial for understanding the structure-function relationship in Fe–S proteins, but it is generally impaired in standard NMR experiments by paramagnetic relaxation enhancement due to the presence of the paramagnetic cluster(s). On the other hand, the availability of systems of different size and stability has, over the years, stimulated NMR spectroscopists to exploit iron-sulfur proteins as paradigmatic cases to develop experiments, models and protocols. Here, the cluster binding properties of human mitoNEET have been investigated by one-dimensional and two-dimensional 1H diamagnetic and paramagnetic NMR, in its oxidized and reduced states. The NMR spectra of both oxidation states of mitoNEET appeared to be significantly different from those reported for previously investigated [Fe2S2]2+/+ proteins. We show how the use of 1D NOE experiments, 13C direct-detected experiments, and the optimization of NMR experiments for paramagnetic systems significantly reduce the blind sphere of the protein around the paramagnetic cluster. The application of this approach provided a detailed description of the unique electronic properties of mitoNEET, that are responsible for its biological function. Indeed, the NMR properties suggested that the specific electronic structure of the cluster possibly drives the functional properties of different [Fe2S2] proteins.

Francesca Camponeschi et al.

Status: final response (author comments only)

Comment types: AC – author | RC – referee | CC – community | EC – editor | CEC – chief editor | : Report abuse
  • RC1: 'Comment on mr-2021-2', Anonymous Referee #1, 22 Jan 2021
    • AC1: 'Reply on RC1', Lucia Banci, 25 Jan 2021
  • RC2: 'Comment on mr-2021-2', Anonymous Referee #2, 15 Feb 2021
    • AC2: 'Reply on RC2', Lucia Banci, 22 Feb 2021
  • EC1: 'Comment on mr-2021-2', Gottfried Otting, 23 Feb 2021

Francesca Camponeschi et al.

Data sets

Paramagnetic tailored experiments for the NMR investigation of reduced and oxidized [2Fe-2S]-mitoNEET Francesca Camponeschi, Angelo Gallo, Mario Piccioli, and Lucia Banci https://doi.org/10.5281/zenodo.4442395

Francesca Camponeschi et al.

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Short summary
The iron-sulfur cluster binding properties of human mitoNEET have been investigated by 1D and 2D 1H paramagnetic NMR spectroscopy. The NMR spectra of both oxidized and reduced mitoNEET are significantly different from those reported previously for others [Fe2S2] proteins. Our findings revealed the unique electronic properties of mitoNEET and suggests that the specific electronic structure of the cluster possibly drives the functional properties of different [Fe2S2] proteins.