The general mitochondrial matrix processing protease from rat liver: structural characterization of the catalytic subunit.
The general mitochondrial matrix processing protease from rat liver: structural characterization of the catalytic subunit.
复制标题
大鼠肝脏的一般线粒体基质加工蛋白酶：催化亚基的结构特征。
DOI：
10.1073/pnas.87.20.7978
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发表时间：
1990
影响因子：
11.1
通讯作者：
Rosenberg,LE
中科院分区：
文献类型：
--
作者：
Kleiber,J;Kalousek,F;Swaroop,M;Rosenberg,LE
将核编码的前体蛋白导入线粒体的关键步骤涉及通过加工线粒体基质中发现的蛋白酶对其氨基末端前导肽进行蛋白水解切割。我们在这里报告的一般基质处理蛋白酶的大鼠肝线粒体的特性。最终的酶制剂由两种多肽组成，一种具有催化活性的55-kDa亚基和一种52-kDa亚基。为了推断55 kDa亚基的完整一级结构，我们首先对其成熟的氨基末端和来自纯蛋白的几个胰蛋白酶肽进行了测序。接下来，使用具有基于这些肽中的两个的序列的混合寡核苷酸引物，我们通过用聚合酶链反应选择性扩增肝RNA来合成部分cDNA探针。然后使用扩增的探针从大鼠肝脏cDNA文库中获得近全长克隆。该cDNA编码508个氨基酸残基，包括氨基末端前导肽的16个残基，其切割位点位于精氨酸残基下游的两个多肽键。成熟的部分具有预测的分子量为55.2 kDa，它与酿酒酵母和粗糙脉孢菌的线粒体加工肽酶具有36%的同一性。一个保守的结构特征是一个假定的，带负电荷的α-螺旋，位于亚基的氨基末端的一半，这个元素可能是重要的识别线粒体前体蛋白的带正电荷的前导肽的特点。
A critical step in the import of nuclear-encoded precursor proteins into mitochondria involves proteolytic cleavage of their amino-terminal leader peptides by processing proteases found in the mitochondrial matrix. We report here the characterization of the general matrix processing protease from rat liver mitochondria. The final enzyme preparation consisted of two polypeptides, a catalytically active 55-kDa subunit and a 52-kDa one. To deduce the complete primary structure of the 55-kDa subunit, we first sequenced its mature amino terminus and several tryptic peptides derived from the pure protein. Next, using mixed oligonucleotide primers that had sequences based on two of these peptides, we synthesized a partial cDNA probe by selective amplification of liver RNA with the polymerase chain reaction. The amplified probe was then used to obtain a nearly full-length clone from a rat liver cDNA library. This cDNA codes for 508 amino acid residues, including 16 residues of an amino-terminal leader peptide, the cleavage site of which is located two polypeptide bonds downstream from an arginine residue. The mature portion has a predicted molecular mass of 55.2 kDa; it shows 36% identity with the mitochondrial processing peptidases of Saccharomyces cerevisiae and Neurospora crassa. A conserved structural feature is a putative, negatively charged alpha-helix, located in the amino-terminal half of the subunit; this element might be important for the recognition of positively charged leader peptides characteristic of mitochondrial precursor proteins.