Fep1 is an iron-responsive GATA-type transcriptional repressor present in numerous fungi. The DNA-binding domain of this protein is characterized by the presence of two zinc fingers of the Cys2-Cys2 type and a Cys-X5-Cys-X8-Cys-X2-Cys motif located between the two zinc fingers, that is involved in binding of a [2Fe-2S] cluster. In this work, biophysical characterization of the DNA-binding domain of Pichia pastoris Fep1 and of the complex of the protein with cognate DNA has been undertaken. The results obtained by analytical ultracentrifugation sedimentation velocity, small-angle X-ray scattering and differential scanning calorimetry indicate that Fep1 is a natively unstructured protein that is able to bind DNA forming 1:1 and 2:1 complexes more compact than the individual partners. Complex formation takes place independently of the presence of a stoichiometric [2Fe-2S] cluster, suggesting that the cluster may play a role in recruiting other protein(s) required for regulation of transcription in response to changes in intracellular iron levels.

Biophysical characterization of the complex between the iron-responsive transcription factor Fep1 and DNA / Miele, A. E.; Cervoni, L.; Le Roy, A.; Cutone, A.; Musci, G.; Ebel, C.; Bonaccorsi di Patti, M. C.. - In: EUROPEAN BIOPHYSICS JOURNAL WITH BIOPHYSICS LETTERS. - ISSN 0175-7571. - 50:3-4(2021), pp. 501-512. [10.1007/s00249-020-01489-y]

Biophysical characterization of the complex between the iron-responsive transcription factor Fep1 and DNA

Miele A. E.;Cervoni L.;Bonaccorsi di Patti M. C.
2021

Abstract

Fep1 is an iron-responsive GATA-type transcriptional repressor present in numerous fungi. The DNA-binding domain of this protein is characterized by the presence of two zinc fingers of the Cys2-Cys2 type and a Cys-X5-Cys-X8-Cys-X2-Cys motif located between the two zinc fingers, that is involved in binding of a [2Fe-2S] cluster. In this work, biophysical characterization of the DNA-binding domain of Pichia pastoris Fep1 and of the complex of the protein with cognate DNA has been undertaken. The results obtained by analytical ultracentrifugation sedimentation velocity, small-angle X-ray scattering and differential scanning calorimetry indicate that Fep1 is a natively unstructured protein that is able to bind DNA forming 1:1 and 2:1 complexes more compact than the individual partners. Complex formation takes place independently of the presence of a stoichiometric [2Fe-2S] cluster, suggesting that the cluster may play a role in recruiting other protein(s) required for regulation of transcription in response to changes in intracellular iron levels.
Analytical ultracentrifugation sedimentation velocity; differential scanning calorimetry; Fep1; fluorescence spectroscopy; iron sulfur cluster; protein-DNA complex; small angle X-ray scattering
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Biophysical characterization of the complex between the iron-responsive transcription factor Fep1 and DNA / Miele, A. E.; Cervoni, L.; Le Roy, A.; Cutone, A.; Musci, G.; Ebel, C.; Bonaccorsi di Patti, M. C.. - In: EUROPEAN BIOPHYSICS JOURNAL WITH BIOPHYSICS LETTERS. - ISSN 0175-7571. - 50:3-4(2021), pp. 501-512. [10.1007/s00249-020-01489-y]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11573/1556816
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