Eusebio A, Friedberg T, Spiess M
Biozentrum, University of Basel, Switzerland.
Exp Cell Res. 1998 May 25;241(1):181-5. doi: 10.1006/excr.1998.4042.
The orientation of signal sequences during insertion into the endoplasmic reticulum membrane is largely determined by the charged residues flanking the apolar domain. Using recombinant and mutant proteins, also length and hydrophobicity of the apolar segment were shown to affect the orientation: translocation of the N-terminus was found to be favored by long hydrophobic sequences, and translocation of the C-terminus, by short ones. Here, we tested the physiological significance of this phenomenon by mutagenesis of the hydrophobic portion of two natural signals with unusual flanking charges. Extending the hydrophobic domain of the short, cleaved Ncyt/Cexo signal of pre-provasopressin-neurophysin II and shortening that of the Nexo/Ccyt signal anchor of microsomal epoxide hydrolase resulted in a significant fraction of polypeptides inserting in the opposite orientation to that of the wild-type proteins. The topogenic contribution of the hydrophobic domain is thus important for the correct and uniform orientation of natural proteins and can explain the behavior of some of the signals with unusual flanking charges.
信号序列插入内质网膜过程中的方向很大程度上由非极性结构域两侧的带电残基决定。利用重组蛋白和突变蛋白，研究还表明非极性片段的长度和疏水性也会影响方向：发现长的疏水序列有利于N端的转运，而短的疏水序列则有利于C端的转运。在此，我们通过对两个具有异常侧翼电荷的天然信号的疏水部分进行诱变，测试了这一现象的生理意义。延长前血管加压素-神经垂体素II的短的、可裂解的Ncyt/Cexo信号的疏水结构域，并缩短微粒体环氧化物水解酶的Nexo/Ccyt信号锚的疏水结构域，导致相当一部分多肽以与野生型蛋白相反的方向插入。因此，疏水结构域的拓扑作用对于天然蛋白的正确和一致方向很重要，并且可以解释一些具有异常侧翼电荷的信号的行为。
