A previously found yeast-mitochondrial protein fraction stabilizing the inactivated complex between mitochondria) ATPase and intrinsic ATPase inhibitor (Hashimoto, T., et al. (1983) J. Biochem. 94, 715-720) was separated into two proteins by high performance liquid chromatography on a cation exchanger. The molecular weights of the factors were estimated to be 9, 000 and 15, 000 daltons by sodium dodecyl sulfate (SDS)-gel electrophoresis. Both factors were required to stabilize a complex of inhibitor and proton-translocating ATPase (F₁F₀-ATPase) either in its purified form or in mitochondrial membranes. On the other hand both factors together could not stabilize a complex of the inhibitor and F₁-ATPase, suggesting that both factors act together with the F₀-portion. The factors also facilitated very efficiently the binding of ATPase inhibitor to F₁F₀-ATPase in the presence of ATP and Mg²⁺. Both the 15, 000 and 9, 000 dalton stabilizing factors were hardly distinguishable from δ- and ε-subunit, respectively, on an SDS-gel electrophoregram, but immuno-diffusion assay showed that neither factor was present in the purified F₁-ATPase containing the δ- and ε-subunit.