OSCP is a subunit of the FA stalk sector of yeast mitochondrial ATP synthase complex. Cells of a null mutant for OSCP, constructed by disruption of the chromosomal ATP5 gene of Saccharomyces cerevisiae, exhibited a high level of genetic instability (petite formation). Study of the effects of ablation of OSCP required the development of a progressive depletion strategy. Introduction of a vector bearing an ATP5 gene cassette under GAL1 transcriptional control into null mutant cells gave rise to a stable yeast strain from which OSCP could be depleted in a controlled manner by manipulation of the level of galactose in the growth medium. Cells progressively depleted of OSCP exhibited properties of cellular respiration indicative of a decline in the functional coupling of the catalytic F1 sector to the proton channel F0 sector (normally linked by FA). Cells depleted of OSCP also exhibited a physical uncoupling of F1 from other subunits of the complex such that other FA subunits and F0 subunit 6 were not recovered in immunoprecipitates of ATP synthase complexes. Thus, OSCP plays a role in the assembly as well as function of the enzyme complex.