Based on available platforms detailing the Saccharomyces cerevisiae mitochondrial proteome and other high-throughput studies, we identified the yeast gene DMO2 as having a profile of genetic and physical interactions that indicate a putative role in mitochondrial respiration. Dmo2p is a homolog to human distal membrane-arm assembly complex protein 1 (DMAC1); both proteins have two conserved cysteines in a Cx₂C motif. Here, we localized Dmo2p in the mitochondrial inner membrane with the conserved cysteines facing the intermembrane space. The respiratory deficiency of dmo2 mutants at 37^oC led to a reduction in cytochrome c oxidase (COX) activity (COX) and in the formation of cytochrome bc₁ complex–COX supercomplexes; dmo2 also has a rapid turnover of Cox2p, the second subunit of the COX complex that harbors the binuclear Cu_A center. Moreover, Dmo2p co-immunoprecipitates with Cox2p and components required for maturation of the Cu_A center, such as Sco1p and Sco2p. Finally, DMO2 overexpression can suppress cox23 respiratory deficiency, a mutant that has impaired mitochondrial copper homeostasis. Mass spectrometry data unveiled the interaction of Dmo2p with different large molecular complexes, including bc₁–COX supercomplexes, the TIM23 machinery, and the ADP/ATP nucleotide translocator. Overall, our data suggest that Dmo2p is required for Cox2p maturation, potentially by aiding proteins involved in copper transport and incorporation into Cox2p

Protein quantification was conducted utilising the MaxQuant label‐free algorithm (LFQ), which employed unique and razor peptides for accurate measurement. A minimum requirement of ≥ 2 ratio counts was set as essential for valid protein quantification.

Mass spectrometry analyses were performed at the Redox Proteomics Core of the Mass Spectrometry Resource at Chemistry Institute, University of São Paulo; the results were processed using the perseus software suite. Data were transformed [log2(x)] and filtered so that for each protein, at least one group (Dmo2‐PC/wild‐type) contained a minimum of 50% valid values. Subsequent steps in perseus included a two‐sample t‐test to compare the means of the label‐free quantitation (LFQ) intensity values between the Dmo2‐PC and wild‐type groups.