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Molecular mechanisms of protein sorting between the mitochondrial outer and inner membranes

线粒体外膜和内膜之间蛋白质分选的分子机制

基本信息

批准号：
63580151
负责人：
HASE Toshiharu
金额：
$ 1.28万
依托单位：
Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1988
资助国家：
日本
起止时间：
1988 至 1989
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-63580151/
关键词：
Mitochondria Membrane proteins Intracellular transport of proteins Cytochrome c_1 チトクロムc_1

项目摘要

Mitochondrial proteins synthesized in the cytoplasm are transported and localized to a specific compartment of the mitochondrion. The intramitochondrial sorting between the outer and inner membranes was investigated using a 70 kDa outer membrane protein and cytochrome c_1 as representative proteins in yeast cells. The presequence of cytochrome c_1, which functions as a signal to the inner membrane was replaced by various lengths of the N-terminal region of the 70 kDa protein. The fused protein with the N-terminal 61 residues(61mC_1) was localized to the outer membrane and those with less than 29 residues to the inner membrane. The latter fusion proteins were shown to be assembled into succinate-eytochrome c reductase without proteolytic removal of the fused region and restore the respiration of a cytochrome c_1 deficient mutant yeast(MH6-16). These date indicate that targeting and anchoring signals of the outer membrane protein localize the inner membrane protein to the outer membrane and that the target sequence alone directs it to the inner membrane.The cytochrome c_1 deficient mutant yeast, MH6-16 transformed with a plasmid encoding 61mC_1 was not able to grow on a non- fermentable medium(glycerol^-), due to location of this protein to the outer membrane. A selection for mutations which restored growth on glycerol was applied to the transformants and yield a class of invitation in the host genome. We are now characterizing this mutation as a mitochondrial protein sorting mutant and a DNA fragment involved in the restoration are currently isolated.

在细胞质中合成的线粒体蛋白被运输并定位到线粒体的特定隔室。以70 kDa的外膜蛋白和细胞色素c_1为代表蛋白，研究了酵母细胞的线粒体内外膜分选。作为内膜信号的细胞色素c_1的序列被70 kDa蛋白的不同长度的n端区域所取代。含有61个n末端残基的融合蛋白（61mC_1）定位于外膜，少于29个残基的融合蛋白定位于内膜。后一种融合蛋白被证明可以组装成琥珀酸-细胞色素c还原酶，而无需蛋白水解去除融合区域，并恢复细胞色素c_1缺陷突变酵母（MH6-16）的呼吸。这些数据表明，外膜蛋白的靶向和锚定信号使内膜蛋白定位于外膜，而单靠靶标序列就能将其导向内膜。用编码61mC_1的质粒转化的细胞色素c_1缺陷突变酵母MH6-16，由于该蛋白位于外膜上，不能在不可发酵的培养基（甘油^-）上生长。对在甘油上恢复生长的突变进行了选择，并在宿主基因组中产生了一类邀请。我们现在将这种突变定性为线粒体蛋白分选突变，目前已分离出参与恢复的DNA片段。