Subunits of the translocon interact with components of the oligosaccharyl transferase complex
Subunits of the translocon interact with components of the oligosaccharyl transferase complex
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DOI：
10.1074/jbc.m502858200
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发表时间：
2005-06-17
影响因子：
4.8
通讯作者：
Lennarz, WJ
中科院分区：
文献类型：
--
作者：
Chavan, M;Yan, AX;Lennarz, WJ
通过转座子在内质网膜上开始易位后，多肽链被寡糖转移酶（OT）酶复合物n -糖基化。易位和n -糖基化是同时发生的事件，预计需要易位和OT复合体并置。为了确定是否有OT复合体和转位子的亚基介导这两个复合体之间的相互作用，我们使用分裂泛素方法在出芽酵母中进行了系统研究。有趣的是，OT亚基Stt3p被发现只与Sec61p相互作用，而另一个OT亚基Ost4p被发现与转座子的所有三个组成部分Sec61p、sh1p和Sss1p相互作用。发现OT亚基Wbp1p与Sec61p和sh1p相互作用非常强，与Sss1p相互作用弱。其他OT亚基，Ost1p， Ost2p和Swp1p被发现与Sec61p和sh1p或Sss1p相互作用。Ost3p与Sec61p和sh1p的相互作用弱，而Ost5p和Ost6p与Sec61p的相互作用非常弱，无法与sh1p或Sss1p相互作用。我们能够通过化学交联技术证实这些分裂泛素的发现。基于我们使用这两种技术的发现，我们得出结论，这两种复合物的结合是通过多种蛋白质-蛋白质接触来稳定的。基于原核Sec复合物晶体结构参数向真核复合物的外推，我们提出了一个理解translocon-OT超复合物组织的工作模型。
Following initiation of translocation across the membrane of the endoplasmic reticulum via the translocon, polypeptide chains are N-glycosylated by the oligosaccharyl transferase (OT) enzyme complex. Translocation and N-glycosylation are concurrent events and would be expected to require juxtaposition of the translocon and the OT complex. To determine whether any of the sub-units of the OT complex and translocon mediate interactions between the two complexes, we initiated a systematic study in budding yeast using the split-ubiquitin approach. Interestingly, the OT subunit Stt3p was found to interact only with Sec61p, whereas another OT subunit, Ost4p, was found to interact with all three components of the translocon, Sec61p, Sbh1p, and Sss1p. The OT subunit Wbp1p was found to interact very strongly with Sec61p and Sbh1p and weakly with Sss1p. Other OT subunits, Ost1p, Ost2p, and Swp1p were found to interact with Sec61p and either Sbh1p or Sss1p. Ost3p exhibited a weak interaction with Sec61p and Sbh1p, whereas Ost5p and Ost6p interacted very weakly with Sec61p and failed to interact with Sbh1p or Sss1p. We were able to confirm these split-ubiquitin findings by a chemical cross-linking technique. Based on our findings using these two techniques, we conclude that the association of these two complexes is stabilized via multiple protein-protein contacts. Based on extrapolation of the structural parameters of the crystal structure of the prokaryotic Sec complex to the eukaryotic complex, we propose a working model to understand the organization of the translocon-OT supercomplex.