Arakawa H, Amaya Y, Mori M
Institute for Medical Genetics, Kumamoto University Medical School.
J Biochem. 1990 Jan;107(1):160-4. doi: 10.1093/oxfordjournals.jbchem.a123001.
Unlike most mitochondrial matrix proteins, the mitochondrial 3-oxoacyl-CoA thiolase [EC 2.3.1.16] is synthesized with no cleavable presequence and possesses information for mitochondrial targeting and import in the mature protein. This mitochondrial thiolase is homologous with the mature portion of peroxisomal 3-oxoacyl-CoA thiolase and acetoacetyl-CoA thiolase [EC 2.3.1.9] of Zoogloea ramigera along the entire sequence. A hybrid gene encoding the NH2-terminal 16 residues (MALLRGVFIVAAKRTP) of the mitochondrial thiolase fused to the mature portion of rat ornithine carbamoyltransferase [EC 2.1.3.3] (lacking its own presequence) was transfected into COS cells, and subcellular localization of the fusion protein was analyzed. Cell fractionation and immunocytochemical analyses showed that the fusion protein was localized in the mitochondria. These results indicate that the NH2-terminal 16 residues of the mitochondrial thiolase function as a noncleavable signal for mitochondrial targeting and import of this enzyme protein. The fusion protein containing the NH2-terminal 14 residues (MSTPSIVIASARTA) of the bacterial thiolase was also localized in the mitochondria. On the other hand, the fusion protein containing the corresponding portion (MQASASDVVVVHGQRTP) of the peroxisomal thiolase appeared not to be localized to the mitochondria. These results show that the import signal of mitochondrial 3-oxoacyl-CoA thiolase originated from the NH2-terminal portion of the ancestral thiolase. The ancestral enzyme might have already possessed a mitochondrial import activity when mitochondria appeared first, or that it might have acquired the import activity during evolution by accumulation of point mutations in the NH2-terminal portion of the enzyme.
与大多数线粒体基质蛋白不同，线粒体3-氧代酰基辅酶A硫解酶[EC 2.3.1.16]在合成时没有可裂解的前导序列，并且在成熟蛋白中含有线粒体靶向和导入的信息。这种线粒体硫解酶在整个序列上与过氧化物酶体3-氧代酰基辅酶A硫解酶的成熟部分以及枝动胶菌的乙酰乙酰辅酶A硫解酶[EC 2.3.1.9]同源。将编码线粒体硫解酶的NH2末端16个残基（MALLRGVFIVAAKRTP）与大鼠鸟氨酸氨甲酰基转移酶[EC 2.1.3.3]的成熟部分（缺乏其自身的前导序列）融合的杂合基因转染到COS细胞中，并分析融合蛋白的亚细胞定位。细胞分级分离和免疫细胞化学分析表明，融合蛋白定位于线粒体中。这些结果表明，线粒体硫解酶的NH2末端16个残基作为该酶蛋白线粒体靶向和导入的不可裂解信号发挥作用。含有细菌硫解酶NH2末端14个残基（MSTPSIVIASARTA）的融合蛋白也定位于线粒体中。另一方面，含有过氧化物酶体硫解酶相应部分（MQASASDVVVVHGQRTP）的融合蛋白似乎没有定位于线粒体。这些结果表明，线粒体3-氧代酰基辅酶A硫解酶的导入信号起源于祖先硫解酶的NH2末端部分。祖先酶可能在最初出现线粒体时就已经具有线粒体导入活性，或者它可能在进化过程中通过该酶NH2末端部分点突变的积累而获得了导入活性。
