Azevedo J E, Nehls U, Eckerskorn C, Heinrich H, Rothe H, Weiss H, Werner S
Institut für Physiologische Chemie, Universität München, Federal Republic of Germany.
Biochem J. 1992 Nov 15;288 ( Pt 1)(Pt 1):29-34. doi: 10.1042/bj2880029.
The 20.9 kDa subunit of NADH:ubiquinone oxidoreductase (complex I) from Neurospora crassa is a nuclear-coded component of the hydrophobic arm of the enzyme. We have determined the primary structure of this subunit by sequencing a full-length cDNA and a cleavage product of the isolated polypeptide. The deduced protein sequence is 189 amino acid residues long and contains a putative membrane-spanning domain. Striking similarity over a 60 amino-acid-residue domain with the M (matrix) protein of para-influenza virus was found. No other relationship with already known sequences could be detected, leaving the function of this subunit in complex I still undefined. The biogenetic pathway of this polypeptide was studied using a mitochondrial import system in vitro. The 20.9 kDa subunit synthesized in vitro is efficiently imported into isolated mitochondria, where it obtains distinct features of the endogenous subunit. Our results suggest that the 20.9 kDa polypeptide is made on cytosolic ribosomes lacking a cleavable targeting sequence, interacts with the mitochondrial outer membrane (in a process that does not require an energized inner membrane), and is imported into mitochondria at contact sites. The 20.9 kDa subunit is then inserted into the inner membrane acquiring a topology similar to that of the already assembled subunit.
粗糙脉孢菌NADH：泛醌氧化还原酶（复合体I）的20.9 kDa亚基是该酶疏水臂的一个核编码组分。我们通过对全长cDNA和分离多肽的一个裂解产物进行测序，确定了该亚基的一级结构。推导的蛋白质序列长度为189个氨基酸残基，包含一个推定的跨膜结构域。发现该序列在一个60个氨基酸残基的区域与副流感病毒的M（基质）蛋白有显著相似性。未检测到与已知序列的其他关系，因此该亚基在复合体I中的功能仍不明确。利用体外线粒体导入系统研究了该多肽的生物发生途径。体外合成的20.9 kDa亚基能有效地导入分离的线粒体，并在其中获得内源性亚基的独特特征。我们的结果表明，20.9 kDa多肽在缺乏可裂解靶向序列的胞质核糖体上合成，与线粒体外膜相互作用（该过程不需要内膜提供能量），并在接触位点导入线粒体。然后，20.9 kDa亚基插入内膜，获得与已组装亚基相似的拓扑结构。
