Interaction of Kar2p and Sls1p is required for efficient co-translational translocation of secreted proteins in the yeast Yarrowia lipolytica
Interaction of Kar2p and Sls1p is required for efficient co-translational translocation of secreted proteins in the yeast Yarrowia lipolytica
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DOI：
10.1074/jbc.273.47.30903
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发表时间：
1998-11-20
影响因子：
4.8
通讯作者：
Gaillardin, C
中科院分区：
文献类型：
--
作者：
Boisramé, A;Kabani, M;Gaillardin, C
酵母解脂耶氏菌是体内研究信号识别颗粒依赖性靶向途径的模式生物。在本报告中，我们定义了增溶条件，并建立了脂质体Y. polytica微粒体的分离程序，以确定与核糖体连接的含有Sec61p的易位孔的数量。与酵母相比，在这种酵母中，70%至80%的Sec61p与核糖体相关。伴侣蛋白Kar2p和内质网腔内的常驻蛋白Sls1p产物与Sec61p群体部分分离，Sls1p可以与Kar2p共免疫沉淀，并且这两种多肽在酵母双杂交系统中直接相互作用。在SLS1编码序列上进行了定点诱变，使我们能够定义Sls1p的功能域。事实上，当这些突变蛋白中的一个表达时，报告蛋白的共翻译易位受到影响。此外，该蛋白已经失去了与Kar2p相互作用的能力，这两种管腔多肽可能因此合作促进分泌蛋白的共翻译易位。
The yeast Yarrowia lipolytica is a model organism for in vivo study of the signal recognition particle-dependent targeting pathway. In this report, we defined solubilization conditions and set up a fractionation procedure of Y. lipolytica microsomes to determine the amounts of Sec61p-containing translocation pores linked to ribosomes, In contrast to Saccharomyces cerevisiae, from 70 to 80% of Sec61p associates with ribosomes in this yeast. The chaperone protein Kar2p and the Sls1p product, a resident protein of the endoplasmic reticulum lumen, partially fractionate with this Sec61p population, Moreover, Sls1p can be co-immunoprecipitated with Kar2p, and the two polypeptides are shown to directly interact in the yeast two-hybrid system. A site-directed mutagenesis was performed on the SLS1 coding sequence that allowed us to define a functional domain in Sls1p, Indeed, co-translational translocation of a reporter protein is affected when one of these mutant proteins is expressed. Moreover, this protein has lost its capacity to interact with Kar2p, and the two lumenal polypeptides might thus cooperate to promote secretory protein co-translational translocation.