Characteristics of Glycine and Malate Oxidation by Pea Leaf Mitochondria: Evidence of Differential Access to NAD and Respiratory Chains

Abstract

The addition of glycine to pea leaf mitochondria oxidizing malate in either state 3 or state 4 led to a marked increase in the rate of O₂ uptake. Malate was also found to stimulate the state 3 and state 4 rates of O₂ uptake with glycine. The increases in state 4 rates were not due to increases in the rate of O₂ uptake via the alternative pathway, as the state 4 rate in the presence of malate or glycine, or both, was not limited by the rate of electron flow between ubiquinone and O₂.

The rotenone-insensitive state 3 rates of O₂ uptake in the presence of malate or glycine, in freshly isolated mitochondria, could be stimulated by the addition of NAD. However, these NAD-limited, rotenone-insensitive, rates could also be stimulated by the addition of a second NAD-linked substrate. Mitochondria could be further depleted of NAD such that the rotenone-insensitive state 3 rate of glycine oxidation was stimulated threefold by added NAD. Even in these NAD-depleted mitochondria, the rate of O₂ uptake in the presence of glycine was found to be markedly stimulated by malate, indicating that the malate-oxidizing enzymes had access to NAD, within these mitochondria, which was unavailable to glycine decarboxylase. These results are not consistent with a uniform distribution of enzymes within mitochondria with access to a common pool of NAD, and suggest that glycine decarboxylase and the malate-oxidizing enzymes may have differential access, both in terms of size and location, to NAD within pea leaf mitochondria.