Demonstration of an unstable variant of pyruvate dehydrogenase protein (E₁) in cultured fibroblasts from a patient with congenital lactic acidemia

The deficiency of pyruvate dehydrogenase enzyme complex causes congenital lactic acidemia and devastating neurologic abnormalities in newborns and children. In the majority of cases, the basic defect appears to be in the pyruvate dehydrogenase (E₁) component, which consists of two subunits, α and β. Whereas some patients are deficient of a single subunit, in other patients both subunits of E₁ are missing. To find out why two proteins were deficient, we investigated the cultured fibroblasts of a female patient who had missing E₁-α and E₁-β protein bands on Western blot. Radiolabeling-immunoprecipita-tion studies with³⁵S-methionine revealed that patient fibroblasts synthesized normal sized precursor E₁-α and E₁-β proteins, which were presumably transported into mitochondria and processed into normal sized mature proteins. However, pulse-chase analysis showed that a- and /î-pro-teins were degraded rapidly compared to normal. Our findings proved that α and β-subunits were synthesized and processed normally but failed to form a stable structure for incorporation into the pyruvate dehydrogenase complex.