Cytochrome mitochondria isolation

Because of the difficulty of isolating the electron transport chain from the rest of the mitochondrion, it is easiest to measure ratios of components (Table 18-3). Cytochromes a, a3, b, cv and c vary from a 1 1 to a 3 1 ratio while flavins, ubiquinone, and nonheme iron occur in relatively larger amounts. The much larger... 

The control of the respiration process and ATP synthesis shifts as the metabolic state of the mitochondria changes. In an isolated mitochondrion, control over the respiration process in state 4 is mainly due to the proton leak through the mitochondrial inner membrane. This type of control decreases from state 4 to state 3, while the control by the adenine nucleotide and the dicarboxylate carriers, cytochrome oxidase, increases. ATP utilizing reactions and transport activities also increase. Therefore, in state 3, most of the control is due to respiratory chain and substrate transport. 

Cytochromes involved in respiration cannot be considered apart from the regulation of the mitochondrion, the organelle described by Altmann in the decade after MacMunn s observation of cytochromes and then by Benda and Michaelis (see reference [8]). Fifty years passed before systematic investigations of the nature of these structures were carried out by the isolation of the particles, the characterization of their chemistry, and the development of a detailed picture of their fine structure. It became clear that the mitochondrion, where cytochrome oxidase (cytochrome aj) is found, is the organelle of respiration [9-11 ], and that the numbers of cristae within a mitochondrion are related to its oxidative capacity [12]. 

The mitochondrial outer membrane is a physical barrier separating the metabolic processes of the cytoplasm from those of the mitochondrion. Integrity of the outer membrane of mitochondria can be determined using a cytochrome c accessibility assay (Douce et al. 1973). Since holocytochrome c cannot diffuse across intact outer membranes, the ratio of the initial rate of succinate icytochrome c oxidore-ductase activity by the mitochondria to that of the same mitochondria after deliberate lysis of their outer membranes is proportional to the extent of damage to the outer membranes incurred during mitochondrial isolation. 

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