Journal of Biomolecular NMR

, Volume 58, Issue 2, pp 123–128 | Cite as

The IR-15N-HSQC-AP experiment: a new tool for NMR spectroscopy of paramagnetic molecules

  • Simone Ciofi-Baffoni
  • Angelo Gallo
  • Riccardo Muzzioli
  • Mario Piccioli


A crucial factor for the understanding of structure-function relationships in metalloproteins is the identification of NMR signals from residues surrounding the metal cofactor. When the latter is paramagnetic, the NMR information in the proximity of the metal center may be scarce, because fast nuclear relaxation quenches signal intensity and coherence transfer efficiency. To identify residues at a short distance from a paramagnetic center, we developed a modified version of the 15N-HSQC experiment where (1) an inversion recovery filter is added prior to HSQC, (2) the INEPT period has been optimized according to fast relaxation of interested spins, (3) the inverse INEPT has been eliminated and signals acquired as antiphase doublets. The experiment has been successfully tested on a human [Fe2S2] protein which is involved in the biogenesis of iron-sulfur proteins. Thirteen HN resonances, unobserved with conventional HSQC experiments, could be identified. The structural arrangement of the protein scaffold in the proximity of the Fe/S cluster is fundamental to comprehend the molecular processes responsible for the transfer of Fe/S groups in the iron-sulfur protein assembly machineries.


Iron-sulfur proteins Paramagnetic NMR 15N-HSQC Pulse sequences Paramagnetic relaxation Anamorsin 



Dr Maciej Mikolajczyk is gratefully acknowledged for the production of CIAPIN1 samples. We gratefully acknowledge the “Programmi di Ricerca di Rilevante Interesse Nazionale” (PRIN) (2009FAKHZT_001), POR CREO FESR 2007-20013—Finding “Farmaci innovativi per malattie neurodegenerative”, POR CREO FESR 2007-20013—Biovax “Vaccini biotecnologici da genomica strutturale” and Ente Cassa di Risparmio di Firenze for financial support. This work was supported by the European Union ESFRI Instruct Core Centre “Centro di Risonanze Magnetiche” (Italy).


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Copyright information

© Springer Science+Business Media Dordrecht 2014

Authors and Affiliations

  • Simone Ciofi-Baffoni
    • 1
  • Angelo Gallo
    • 1
  • Riccardo Muzzioli
    • 1
  • Mario Piccioli
    • 1
  1. 1.Magnetic Resonance Center and Department of ChemistryUniversity of FlorenceSesto FiorentinoItaly

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