ATP-dependent proteolysis in mitochondria

我们研究了线粒体中atp依赖性的蛋白水解系统。虽然动物线粒体中存在一些atp依赖的蛋白酶，但我们的研究重点是atp依赖的LON蛋白酶。我们从牛肾上腺皮质中纯化了蛋白酶并对其进行了表征。在本研究中，我们发现酶可以受到多肽或蛋白质的刺激，而这种刺激是由于刺激酶释放ADP（反应产物之一）。我们测定了编码蛋白酶的cDNA的核苷酸序列。cDNA的开放阅读框包含960个氨基酸，包括前置序列。为了精确确定前序列的长度，我们测定了成熟酶的n端氨基酸序列。我们发现该酶具有异构n端，从Trp-69和Asn-73之间的氨基酸开始。为了确定该前序列是否包含转运蛋白进入线粒体的信息，我们在中国仓鼠卵巢细胞中表达了由该前序列和绿色荧光蛋白（GFP）组成的融合蛋白，发现绿色荧光被纳入颗粒中，这些颗粒被认为是线粒体。

We have investigated about ATP-dependent proteolytic system in mitochondria. Although there are some ATP-dependent proteases in animal mitochondria, we focused our studies on ATP-dependent LON protease. We purified the protease from bovine adrenal cortex and characterized it. In the present research we found that the enzyme was stimulated by peptides or proteins and that the stimulation was due to the stimulation of release of ADP (one of the reaction products) from the enzyme. We determined nucleotide sequence of the cDNA encoding the protease. The open reading frame of the cDNA contained 960 amino acids including presequence. To determine the precise length of the presequence we determined N-terminal amino acid sequence of the mature enzyme. We revealed that the enzyme has heterogeneous N-terminal starting from an amino acid between Trp-69 and Asn-73. To determine whether the presequence contained the information for transporting the protein into mitochondria, we expressed a fusion protein composed of the presequence and Green Fluorescent Protein (GFP) in Chinese hamster ovary cells and found that the green fluorescence was incorporated into granules which were assumed to be mitochondria.