Changes in the electron transport chain of pea leaf mitochondria metabolizing malate

Pea leaf mitochondria showed complex kinetics for malate metabolism. O2 uptake increased as malate concentration increased from 0 to 10 mM, reached a plateau between 10 and 20 mM malate, and then increased again up to 40 mM malate. Analysis of the products of malate oxidation by high-performance liquid chromatography revealed that the first phase of O2 uptake coincided with the synthesis of both pyruvate and oxalacetate (OAA) while the second phase of O2 uptake at higher malate levels usually occurred with a large increase in OAA formation. The biphasic response in O2 uptake and the changing ratios of pyruvate and OAA synthesis did not appear to be the direct result of the differing Km values of malate dehydrogenase and malic enzyme. Rather, they resulted from thermodynamic properties of these two malate oxidases and the kinetics of the two NADH dehydrogenases found in plant mitochondria. At low malate concentrations the rotenone-sensitive NADH dehydrogenase was active and could accept electrons from both malate oxidases. This NADH dehydrogenase became saturated at about 10 mM malate. At higher malate concentrations the rotenone-insensitive NADH dehydrogenase was increasingly important and its increased electron transport capacity was best exploited by malate dehydrogenase. At the higher malate concentrations an increasing portion of the electrons from malate reduce O2 through the alternative oxidase. Although this coincided with the second phase of malate-dependent O2 uptake it was not required for this phase to be seen.