The enzymic hydrolysis of adenosine triphosphate by liver mitochondria. 1. Activities at different pH values

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Abstract
Normal liver mitochondria hydrolyze adenosine triphosphate (ATP) slowly at all pH values between 4.5 and 8.5. The addition of an uncoupling agent such as 2,4-dinitrophenol produces a marked stimulation of the ATPase activity between pH 6 and 8.5. However, the maximal enzymic activity is only attained when the mitochondrial structure has been disintegrated, e. g. by repeated freezing and thawing. The pH-activity curves obtained with mitochondrial fragments or with normal mitochondria in the presence of 2,4-dinitrophenol appear to represent 4 superimposed optimum pH values which are located in the regions of pH 6.3, 7.4, 8.5 and 9.4. This is interpreted in terms of the presence in mitochondria of 4 different ATPase systems. All of these hydrolytic enzymes seem to be relatively specific for ATP. The 9.4 enzyme is the only one which is appreciably active in freshly prepared liver mitochondria, in the presence of ethylenediaminetetraacetate and absence of uncoupling agents. This activity is not due to irreversible structural damage, brought about by the high pH. This enzyme is not stimulated by 2,4-dinitrophenol but is activated by ageing followed by freezing and thawing. The 8.5 enzyme is stimulated by 2,4-dinitrophenol but requires a higher concentration than the 7.4 and 6.3 enzymes for full activation. It is also activated by damaging the mitochondria. The 7.4 enzyme is stimulated by 2,4-dinitrophenol, by mechanical damage to the mitochondria and by removal of ethylenediaminetetra-acetate from the reaction medium. The 6.3 enzyme is the only one clearly activated by 10-5 [image] -2,4-dinitrophenol, and is also fully activated by ageing the mitochondria. The 6.3, 7.4 and 8.5 enzymes, which are stimulated by 2,4-dinitrophenol, may be related to the 3 steps in the respiratory chain which are coupled with phosphorylation.