Isolation and Oxidative Properties of Intact Mitochondria from the Leaves of Sedum praealtum

A procedure is described for preparing intact mitochondria from leaves of Sedum praealtum D.C., a plant showing Crassulacean acid metabolism. These mitochondria oxidized malate, pyruvate, alpha-ketoglutarate, succinate, NADH, NADPH, and isocitrate with good respiratory control and ADP/O ratios better than those observed in mitochondria from other photosynthetic tissues.Malate oxidation was very resistant to inhibition by rotenone. Glycine oxidation was very slow with poor respiratory control and was resistant to rotenone inhibition. Antimycin A completely inhibited the oxidation of both NADH and NADPH. The oxidation of isocitrate, malate, succinate, and alpha-ketoglutarate was partially inhibited by antimycin A and cyanide. Overall rates of substrate oxidation were slow on a protein basis, but purification of the mitochondrial preparations on a linear sucrose gradient removed a large amount of nonmitochondrial protein. The original mitochondrial preparations contained little glycolate oxidase activity, and most of this activity was removed by the sucrose gradient.