ラットジヒドロリポアミドデヒドロゲナーゼ前駆体のin vitro合成とそのミトコンドリアマトリックスへの移行

概要

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Rat dihydrolipoamide dehydrogenase (DLDH) precursor was synthesized in a nuclease treated rabbit reticulocyte lysate system using rat liver polysomal RNA. Molecular mass of the precursor was estimated to be approximately 59kDa on SDS-PAGE. The precursor was adsorbed on DE-32, and it was eluted in a single form on electric charge by NaCl gradient. After incubation of the precursor with isolated rat liver mitochondria, [^<35>S]-DLDH, which was solubilized from the mitochondria, was resolved into two forms on DE 32 under the condition employed. Each molecular mass of two forms of DLDH separated by DE-32 was estimated to be approximately 55kDa on SDS-PAGE. Since the decrease of the mass of the DLDH precursor corresponded to cleavage of its mitochondria-targeting signal at the mitochondrial matrix, this result indicates the DLDH precursor had entered to the mitochondrial matrix. Thus the present study strongly supports the conclusions that the DLDH precursor is synthesized in the single form on the molecular mass and the electric charge, and the multiplicity of electric charge of the mature DLDH resulted from translocation of the DLDH precursor into the mitochondrial matrix.