Iron–sulfur cluster redox chemistry and dimer dissociation in the outer mitochondrial membrane protein, mitoNEET

• Published in: Journal of biological inorganic chemistry Vol. 30; no. 1; pp. 3 - 11
• Main Authors: Chaudhry, Kanita A., Rajanayake, Krishani K., Carroll, Richard T., Isailovic, Dragan, Funk, Max O.
• Format: Journal Article
• Language: English
• Published: Cham Springer International Publishing 01.02.2025; Springer Nature B.V
• Subjects: Ammonium; Biochemistry; Biomedical and Life Sciences; Chemical reduction; Comment; Cytosol; Diabetes mellitus; Dimerization; Dithionite; Glutathione; Humans; Iron sulfides; Iron-Sulfur Proteins - chemistry; Iron-Sulfur Proteins - metabolism; Life Sciences; Mass spectroscopy; Microbiology; Mitochondria; Mitochondrial Membranes - chemistry; Mitochondrial Membranes - metabolism; Mitochondrial Proteins - chemistry; Mitochondrial Proteins - metabolism; Oxidation-Reduction; Pioglitazone; Protein Multimerization; Proteins; Redox properties; Sodium; Sulfur; Sulfur - chemistry; Redox chemistry; Dithionite; Dissociation; mitoNEET; Electrospray ionization mass spectrometry (ESI–MS); Dimerization; Iron–sulfur cluster
• ISSN: 1432-1327; 0949-8257; 1432-1327
• DOI: 10.1007/s00775-024-02093-7

The outer mitochondrial membrane protein known as mitoNEET was discovered when it was labeled by a photoaffinity derivative of the anti-diabetes medication, pioglitazone. The biological role for mitoNEET and its specific mechanism for achieving this remains an active subject for research. There is accumulating evidence suggesting that mitoNEET could be a component of mitochondrial FeS cofactor biogenesis. The protein was composed of an N-terminal membrane associated domain and a C-terminal domain oriented to the cytosol. The cytosolic domain was an iron–sulfur (2Fe–2S) metalloprotein with a rare 3Cys/1His coordination environment. It was previously reported that mitoNEET formed dimers that were remarkably sensitive to pH, likely a consequence of the protonation of the single His-iron ligand. The hypothesis pursued in the research reported here was that perhaps the dissociation of mitoNEET was also sensitive to the redox state of the iron sulfur cluster. To use native electrospray ionization mass spectrometry (ESI–MS) to monitor the reduction reaction ammonium dithionite was envisioned as the appropriate reagent to avoid sodium ion adduct formation from sodium dithionite. The preparation of ammonium dithionite was updated and the compound had the same properties as the sodium salt with redox dyes and the oxidized form of glutathione. The dissociation of mitoNEET treated with ammonium dithionite anaerobically was readily evident as ammonium dithionite was found to be compatible with redox chemistry evaluated by native ESI–MS. Graphical abstract