Abstract: Cox11 is an intrinsic mitochondrial membrane protein essential for the assembly of an active cytochrome c oxidase complex. Cox11 is tethered to the mitochondrial inner membrane by a single transmembrane helix. Domain mapping was carried to determine the functional segments of the Cox11 protein. The C-terminal 189 residue Cu(I) binding domain is shown to be exposed within the inner mitochondrial membrane space (IMS). This orientation was demonstrated by the proteolytic susceptibility of a C-terminal Myc epitope tag in mitoplasts but not in intact mitochondria. Fusion of the N-terminus of Cox11 to a matrix ribosomal protein Rsm22 results in a functional protein capable of suppressing the respiratory defect of both Dcox11 and Drsm22 cells. The functionality of the fusion protein suggests that the Cox11 N-terminus projects into the matrix. The fusion of C-terminal fusion of Cox11 to Rsm22 resembles a naturally occurring fusion of Cox11 in Schizosaccharomyces pombe to a sequence homologous to Saccharomyces cerevisiae Rsm22. Studies on a series of SCO1/COX11 chimeras reveal that the matrix domain of Cox11 lacks a specific function, whereas the Cu(I) binding/donating function requires the yeast Cox11 sequence. The Cu(I) binding domain from human Cox11 cannot functionally replace the yeast sequence. The Cu domain of Cox11 may be an important docking motif for Cox1 or a Cox1 associated protein.