Provisions of reductant for the hydroxypyruvate to glycerate conversion in leaf peroxisomes : a critical evaluation of the proposed malate/aspartate shuttle

A series of experiments, with Secale cereale and Triticum aestivum var Argee, to evaluate critically the ability of a malate/aspartate shuttle to provide reducing equivalents to drive hydroxypyruvate reduction to glycerate led to the conclusion that the shuttle, as previously envisioned, does not supply NADH to the peroxisomal matrix. First, analysis of coupled malate dehydrogenase and glutamate-oxaloacetate transaminase activities in the directions required for intraperoxisomal NADH generation indicated that the peroxisomal enzyme activities were insufficient to account for necessary rates of photorespiratory carbon flux. Second, although the peroxisomal isozyme of malate dehydrogenase comprised a substantial portion (40%) of total cellular activity, less than 7% of the cellular glutamate-oxaloacetate transmaminase activity was associated with the peroxisomes. Third, a peroxisomal extract was able to reduce added NAD only slowly upon addition of malate and glutamate. The rate of NAD reduction was greatly enhanced in the presence of exogenously added glutamateoxaloacetate transaminase. Finally, intact peroxisomes were unable to reduce hydroxypyruvate to glycerate when supplied with malate and glutamate in the absence of exogenously added pyridine nucleotides, although they readily reduced hydroxypyruvate when exogenous pyridine nucleotides were supplied. Three alternative mechanisms, which are in agreement with observed data and which could serve to supply the reducing power to the peroxisomal matrix, are discussed.