Nanobodies as tools for studying human frataxin biology
Iron-sulfur clusters are essential cofactors for the accurate cellular function of many proteins. In eukaryotic cells, the biogenesis of most iron-sulfur clusters occurs in the mitochondria and involves the action of the Cys desulfurase supercomplex, which is activated by the protein frataxin (FXN). The decrease of FXN expression and/or function results in Friedreich's ataxia (FRDA).In this work, several nanobodies specific to human FXN were selected via phage display, demonstrating a wide range of effects on Cys desulfurase activity and a strong interaction with FXN. Nanobody interaction stabilized wild-type and FRDA-related FXN variants in vitro. FXN-nanobody complexes were characterized by NMR, SAXS, and X-ray crystallography. Additionally, Nanobody expression was studied in human cells. The subcellular localization, direct interaction with FXN by in situ proximity ligation assay, effect on cell viability, Fe-S-dependent enzymatic activities, and oxygen consumption rates were analyzed. Significantly, nanobody expression did not alter these key metabolic variables, suggesting that the interaction with FXN did not disrupt the pathway.As a whole, our results suggest that nanobodies can serve as binding partners for mitochondrial FXN. However, the specific effect of the nanobodies on the conformational stability of FRDA-related FXN variants in cells should be investigated.
Authors
- Natalia Fernández (ORCID: https://orcid.org/0000-0001-9170-2101)
- J. Santos
- Viviana Parreño (ORCID: https://orcid.org/0000-0003-0385-5592)
- J.A. Hermoso (ORCID: https://orcid.org/0000-0002-1862-8950)
- María Florencia Pavan (ORCID: https://orcid.org/0009-0007-3646-5749)
- Antonella Vila (ORCID: https://orcid.org/0000-0002-1294-5824)
- Martín E. Noguera (ORCID: https://orcid.org/0000-0001-5868-3834)
- Rafael Molina (ORCID: https://orcid.org/0000-0002-6443-4989)
- Alba Garay-Alvarez (ORCID: https://orcid.org/0009-0001-6544-7327)
- Julian Grossi
- Martin Aran
- Augusto E. García
- Lorena Itatí Ibañez
- Inés G. Muñoz
- Hernán G. Gentili
- Marina Bok
- Naira Antonia Rodríguez
- María Florencia Pignataro
Institutions
- Consejo Superior de Investigaciones Científicas (ES)
- Consejo Nacional de Investigaciones Científicas y Técnicas (AR)
- Universidad de Flores (AR)
- Universidad de Buenos Aires (AR)
- Fundación Instituto Leloir (AR)
- National Institute of Industrial Technology (AR)
- Spanish National Cancer Research Centre (ES)
- Centro Científico Tecnológico - San Juan (AR)
- Instituto Nacional de Tecnologia (BR)
- Instituto de Química y Fisicoquímica Biológicas (AR)
- National Agricultural Technology Institute (AR)
- Instituto de Biotecnología y Biología Molecular (AR)
- Fundación Ciencias Exactas y Naturales (AR)
Publication Details
- Journal
- Communications Biology
- Published
- 2026-01-03
- DOI
- https://doi.org/10.1038/s42003-025-09458-x
- Primary Topic
- Genetic Neurodegenerative Diseases
- Type
- article
- Field-Weighted Citation Impact
- 0.00
Funders
- Friedreich's Ataxia Research Alliance
- Consejo Nacional de Investigaciones Científicas y Técnicas
- Ministerio de Ciencia e Innovación
- Universidad de Buenos Aires
- Secretaría de Ciencia y Técnica, Universidad de Buenos Aires