ROLE OF CARDIOLIPIN ON THE PURIFICATION OF RECONSTITUTIVELY ACTIVE MITOCHONDRIAL PHOSPHATE CARRIER

Publisher Summary This chapter discusses the role of cardiolipin on the purification of reconstitutively active mitochondrial phosphate carrier. The basic procedure for purifying the mitochondrial Pi carrier in active form and in large amounts includes solubilization of the mitochondria in nonionic detergents and chromatography on hydroxylapatite (HTP). The role of cardiolipin on the purification of reconstitutively active Pi carrier has been reinvestigated by measuring the Pi transport activity in both mitochondrial extracts and HTP eluates. Cardiolipin has several effects—it protects the Pi carrier against inactivation by Triton, it decreases the solubilization of mitochondrial protein and of Pi carrier, it activates the phosphate carrier when added to mitochondrial extracts, and it is needed for elution of the Pi carrier from HTP. The latter effect is of primary importance for the preparation of highly active purified Pi carrier. The mitochondrial Pi carrier has an essential requirement for DPG. The inhibition of the Pi carrier by Triton can be prevented or partly released by DPG. It is likely that Triton inhibits the Pi carrier by removing the endogenous DPG from the carrier protein.