While its biological function remains unclear, the 3-Cys, 1-His ligated human [2Fe-2S] cluster containing protein mitoNEET is of interest due to its interaction with the anti-diabetes drug pioglitazone. The mitoNEET [2Fe-2S] cluster demonstrates proton coupled-electron transfer (PCET) and marked cluster instability which have both been linked to the single His-ligand. Highly conserved hydrogen bonding networks, which include the His-87 ligand, exist around the [2Fe-2S] cluster. Through a series of site-directed mutations the PCET of the cluster has been examined, demonstrating that multiple sites of protonation exist in addition to the His-ligand, which can influence redox potential. The mutations also demonstr... More
While its biological function remains unclear, the 3-Cys, 1-His ligated human [2Fe-2S] cluster containing protein mitoNEET is of interest due to its interaction with the anti-diabetes drug pioglitazone. The mitoNEET [2Fe-2S] cluster demonstrates proton coupled-electron transfer (PCET) and marked cluster instability which have both been linked to the single His-ligand. Highly conserved hydrogen bonding networks, which include the His-87 ligand, exist around the [2Fe-2S] cluster. Through a series of site-directed mutations the PCET of the cluster has been examined, demonstrating that multiple sites of protonation exist in addition to the His-ligand, which can influence redox potential. The mutations also demonstrate that while replacement of the His-ligand with cysteine results in a stable cluster, the removal of Lys-55 also greatly stabilizes the cluster. We have also noted for the first time that the oxidation state of the cluster controls stability; the reduced cluster is stable, while the oxidized one is much more labile. Finally, it is shown that upon cluster loss the mitoNEET protein structure becomes less stable, while upon in vitro reconstitution both cluster and secondary structure are recovered. Recently two other proteins have been identified with a 3-Cys(sulfur) 1-His motif, IscR and Grx3/4-Fra2, both of which are sensors of iron and redox homeostatsis. These results lead to a model in which mitoNEET could sense cellular oxidation state and proton concentration and respond through cluster loss and unfolding.
