Functional investigation of an universally conserved leucine residue in subunit a of ATP synthase targeted by the pathogenic m.9176 T>G mutation

• Published in: Biochimica et biophysica acta. Bioenergetics 2019-01, Vol.1860 (1), p.52-59
• Main Authors: Kucharczyk, Roza, Dautant, Alain, Godard, François, Tribouillard-Tanvier, Déborah, di Rago, Jean-Paul
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - biosynthesis; Conserved Sequence; Humans; Leigh Disease - etiology; Leucine - physiology; Life Sciences; Mitochondrial Myopathies - etiology; Mitochondrial Proton-Translocating ATPases - chemistry; Mitochondrial Proton-Translocating ATPases - genetics; Mutation; Protein Subunits; Retinitis Pigmentosa - etiology; Saccharomyces cerevisiae Proteins - genetics; Structure-Activity Relationship
• ISSN: 0005-2728; 1879-2650
• DOI: 10.1016/j.bbabio.2018.11.005

Protons are transported from the mitochondrial matrix to the intermembrane space of mitochondria during the transfer of electrons to oxygen and shuttled back to the matrix by the a subunit and a ring of identical c subunits across the membrane domain (FO) of ATP synthase, which is coupled to ATP synthesis. A mutation (m.9176 T > G) of the mitochondrial ATP6 gene that replaces an universally conserved leucine residue into arginine at amino acid position 217 of human subunit a (aL217R) has been associated to NARP (Neuropathy, Ataxia and Retinitis Pigmentosa) and MILS (Maternally Inherited Leigh's Syndrome) diseases. We previously showed that an equivalent thereof in Saccharomyces cerevisiae (aL237R) severely impairs subunit a assembly/stability and decreases by >90% the rate of mitochondrial ATP synthesis. Herein we identified three spontaneous first-site intragenic suppressors (aR237M, aR237T and aR237S) that fully restore ATP synthase assembly. However, mitochondrial ATP synthesis rate was only partially recovered (40–50% vs wild type yeast). In light of recently described high-resolution yeast ATP synthase structures, the detrimental consequences of the aL237R change can be explained by steric and electrostatic hindrance with the universally conserved subunit a arginine residue (aR176) that is essential to FO activity. aL237 together with three other nearby hydrophobic residues have been proposed to prevent ion shortage between two physically separated hydrophilic pockets within the FO. Our results suggest that aL237 favors subunit c-ring rotation by optimizing electrostatic interaction between aR176 and an acidic residue in subunit c (cE59) known to be essential also to the activity of FO. •Molecular basis of diseases induced by the m.9176 T > G mutation•Intragenic suppressors of the m.9176 T > G mutation in yeast•Role of the leucine residue at position 237 of yeast subunit a