The two redox states of the human NEET proteins’ [2Fe–2S] clusters

Ke Zuo, Henri Baptiste Marjault, Kara L. Bren, Giulia Rossetti, Rachel Nechushtai*, Paolo Carloni*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

5 Scopus citations


The NEET proteins constitute a unique class of [2Fe–2S] proteins. The metal ions bind to three cysteines and one histidine. The proteins’ clusters exist in two redox states; the oxidized protein (containing two FeIII ions) can transfer the cluster to apo-acceptor protein(s), while the reduced form (containing one ferrous ion) remains bound to the protein frame. Here, we perform in silico and in vitro studies on human NEET proteins in both reduced and oxidized forms. Quantum chemical calculations on all available human NEET proteins structures suggest that reducing the cluster weakens the Fe–NHis and Fe–SCys bonds, similar to what is seen in other Fe–S proteins (e.g., ferredoxin and Rieske protein). We further show that the extra electron in the [2Fe–2S]+ clusters of one of the NEET proteins (mNT) is localized on the His-bound iron ion, consistently with our previous spectroscopic studies. Kinetic measurements demonstrate that the mNT [2Fe–2S]+ is released only by an increase in temperature. Thus, the reduced state of human NEET proteins [2Fe–2S] cluster is kinetically inert. This previously unrecognized kinetic inertness of the reduced state, along with the reactivity of the oxidized state, is unique across all [2Fe–2S] proteins. Finally, using a coevolutionary analysis, along with molecular dynamics simulations, we provide insight on the observed allostery between the loop L2 and the cluster region. Specifically, we show that W75, R76, K78, K79, F82 and G85 in the latter region share similar allosteric characteristics in both redox states. Graphic abstract: [Figure not available: see fulltext.].

Original languageAmerican English
Pages (from-to)763-774
Number of pages12
JournalJournal of Biological Inorganic Chemistry
Issue number7
StatePublished - Oct 2021

Bibliographical note

Publisher Copyright:
© 2021, The Author(s).


  • Ab initio calculations
  • Evolutionary analysis
  • Human NEET proteins
  • Iron–sulfur cluster lability
  • Kinetic measurements


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