Regulation of pyruvate dehydrogenase in isolated rat liver mitochondria. Effects of octanoate, oxidation-reduction state, and adenosine triphosphate to adenosine diphosphate ratio.

Factors which influence the distribution of pyruvate dehydrogenase between its active, unphosphorylated form (PDHB) and its inactive, phosphorylated form (PDHb) have been examined in isolated rat liver mitochondria.A rapid freezing method was developed for the extraction of pyruvate dehydrogenase from incubated mitochondria which prevented interconversions between PDH, and PDHb which normally occur when mitochondria are collected by centrifugal methods.The intramitochondrial ATP: ADP ratio was varied over a loo-fold range by the addition of dinitrophenol, oligomycin, or both substances to mitochondria oxidizing 2-0x0glutarate.PDH, activity was found to be inversely proportional to the intramitochondrial ATP:ADP ratio but was not closely correlated with the extramitochondrial adenine nucleotide levels.When mitochondria were incubated in State 4 with succinate and rotenone, the addition of pyruvate increased PDH, activity more than lo-fold without appreciably altering the mitochondrial ATP:ADP ratio.These observations are most readily explained by the known inhibitory effects of pyruvate and ADP on PDH, kinase.PDH, activity could be maintained at a high level by incubating mitochondria in a condition resembling State 3 by the addition of succinate, glucose, and hexokinase.The further addition of octanoate reduced PDH, activity by 60 % without appreciably altering the ATP:ADP ratio.Rotenone had a similar effect.When added in the presence of octanoate, rotenone further decreased PDH, activity whereas 4-pentenoate led to an increase in activity.The effects of OCtanoate on PDH, activity were not seen when mitochondria were incubated in the presence of high levels of pyruvate, though pyruvate oxidation was still diminished by over 50%.The data suggest that octanoate addition favors the PDH,