Mutation of the His ligand in mitoNEET stabilizes the 2Fe-2S cluster despite conformational heterogeneity in the ligand environment

Andrea R. Conlan, Mark L. Paddock, Christina Homer, Herbert L. Axelrod, Aina E. Cohen, Edward C. Abresch, John A. Zuris, Rachel Nechushtai, Patricia A. Jennings

Research output: Contribution to journalArticlepeer-review


MitoNEET is the only identified Fe-S protein localized to the outer mitochondrial membrane and a 1.5 Å resolution X-ray analysis has revealed a unique structure [Paddock et al. (2007), Proc. Natl Acad. Sci. USA, 104, 14342-14347]. The 2Fe-2S cluster is bound with a 3Cys-1His coordination which defines a new class of 2Fe-2S proteins. The hallmark feature of this class is the single noncysteine ligand His87, which when replaced by Cys decreases the redox potential (E m) by 300 mV and increases the stability of the cluster by around sixfold. Unexpectedly, the pH dependence of the lifetime of the 2Fe-2S cluster remains the same as in the wild-type protein. Here, the crystal structure of H87C mitoNEET was determined to 1.7 Å resolution (R factor = 18%) to investigate the structural basis of the changes in the properties of the 2Fe-2S cluster. In comparison to the wild type, structural changes are localized to the immediate vicinity of the cluster-binding region. Despite the increased stability, Cys87 displays two distinct conformations, with distances of 2.3 and 3.2 Å between the S and the outer Fe of the 2Fe-2S cluster. In addition, Lys55 exhibits multiple conformations in the H87C mutant protein. The structure and distinct characteristics of the H87C mutant provide a framework for further studies investigating the effects of mutation on the properties of the 2Fe-2S cluster in this new class of proteins.

Original languageAmerican English
Pages (from-to)516-523
Number of pages8
JournalActa Crystallographica Section D: Biological Crystallography
Issue number6
StatePublished - Jun 2011


  • 2Fe-2S clusters
  • CISD1
  • diabetes
  • mitochondrial outer membrane
  • pioglitazone

All Science Journal Classification (ASJC) codes

  • Structural Biology


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