LATENT MEMBRANE PERTURBATION ACTIVITY OF A MITOCHONDRIAL PRECURSOR PROTEIN IS EXPOSED BY UNFOLDING
LATENT MEMBRANE PERTURBATION ACTIVITY OF A MITOCHONDRIAL PRECURSOR PROTEIN IS EXPOSED BY UNFOLDING
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DOI：
10.1002/j.1460-2075.1988.tb02925.x
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发表时间：
1988-04-01
期刊：
影响因子：
11.4
通讯作者：
SCHATZ, G
中科院分区：
文献类型：
--
作者：
ENDO, T;SCHATZ, G
我们纯化了毫克量的可进口线粒体前体蛋白[酵母细胞色素氧化酶亚基IV融合到小鼠二氢叶酸还原酶（DHFR）的前体]。这使得首次对前体蛋白的构象及其与脂质膜的相互作用进行详细的研究成为可能。前体蛋白在二级结构（通过CD光谱测量）和对尿素的稳定性（通过色氨酸荧光和酶活性测量）方面与真实小鼠DHFR非常相似。有了这个前体蛋白，前体序列就不会显著改变附着的“乘客蛋白”的折叠。与相应的前体肽相比，天然前体仅表现出较弱的破坏具有低摩尔率带负电荷脂质的囊泡的能力，这表明客运蛋白掩盖了前体的两亲性。通过增加囊泡中带负电荷的脂质含量或在5 M尿素中变性前驱体，可以大大增强前驱体的膜扰动特性。与富含酸性磷脂的囊泡的相互作用伴随着前体的部分展开，这表明这种构象变化也可能与前体与线粒体膜的相互作用有关。
We have purified milligram amounts of an importable mitochondrial precursor protein [the presequence of yeast cytochrome oxidase subunit IV fused to mouse dihydrofolate reductase (DHFR)]. This has made it possible, for the first time, to perform detailed studies on the conformation of a precursor protein and its interaction with lipid membranes. The precursor protein closely resembled authentic mouse DHFR with respect to secondary structure (measured by CD spectra) and stability towards urea (measured by tryptophan fluorescence and enzyme activity). With this precursor protein, the presequence thus does not significantly alter the folding of the attached ''passenger protein''. In contrast to the corresponding presequence peptide, the native precursor exhibited only weak ability to disrupt vesicles with a low mol% of negatively charged lipids, suggesting that the passenger protein masks the amphiphilic properties of the presequence. The membrane-perturbing properties of the precursor were greatly enhanced by increasing the vesicles'' content negatively charged lipid or by denaturing the precursor in 5 M urea. Interaction with vesicles rich in acidic phospholipid was accompanied by partial unfolding of the precursor, suggesting that such a conformational change may also be involved in the interaction of the precursor with the mitochondrial membranes.