Vitamin E deficiency in the rat. Cytoplasmic factors required for suppression of mitochondrial respiratory decline

Liver cytoplasm from either vitamin E-deficient or normal rats has been shown to suppress mitochondrial respiratory decline induced by microsomes in vitro. The present study is an attempt to identify cytoplasmic factor(s) responsible for this suppression. The effect of cytoplasm from both groups of animals could be removed by trichloroacetate precipitation, boiling, and dialysis against 0.3 M mannitol-o.1 mM Tris buffer, pH 7.4, but not by aging or extraction with isooctane. The addition of oxalacetate of alpha-ketoglutarate plus asparate to cytolplasm potentiated the suppressant effect. Dialyzed cytoplasm was not effective in depressing the respiratory decline. However, the addition of NAD, osalacetate, or alpha-ketoglutarate plus aspartate to dialyzed cytoplasm restored its ability to depress the respiratory decline. When oxalacetate or alpha-ketoglutarate plus aspartate was added to fresh cytoplasm, followed by dialysis, the effect of these compounds was not seen. These results suggest that cytoplasmic factor(s) required for suppression of the respiratory decline were enzymes of the malate shuttle and transamination, which are heat labile and trichloroacetate precipitable, plus the dialyzable metabolites that are associated with these enzymes in the generation of NAD in mitochondria.