Wittke Sandra, Dünnwald Martin, Albertsen Markus, Johnsson Nils
Max-Delbrück-Laboratorium, 50829 Cologne, Germany.
Mol Biol Cell. 2002 Jul;13(7):2223-32. doi: 10.1091/mbc.01-10-0518.
Ssh1p of Saccharomyces cerevisiae is related in sequence to Sec61p, a general receptor for signal sequences and the major subunit of the channel that guides proteins across the membrane of the endoplasmic reticulum. The split-ubiquitin technique was used to determine whether Ssh1p serves as an additional receptor for signal sequences in vivo. We measured the interactions between the N(ub)-labeled Ssh1p and C(ub)-translocation substrates bearing four different signal sequences. The so-determined interaction profile of Ssh1p was compared with the signal sequence interaction profile of the correspondingly modified N(ub)-Sec61p. The assay reveals interactions of Ssh1p with the signal sequences of Kar2p and invertase, whereas Sec61p additionally interacts with the signal sequences of Mfalpha1 and carboxypeptidase Y. The measured physical proximity between Ssh1p and the beta-subunit of the signal sequence recognition particle receptor confirms our hypothesis that Ssh1p is directly involved in the cotranslational translocation of proteins across the membrane of the endoplasmic reticulum.
酿酒酵母的Ssh1p在序列上与Sec61p相关，Sec61p是信号序列的通用受体，也是引导蛋白质穿过内质网膜的通道的主要亚基。采用分裂泛素技术来确定Ssh1p在体内是否作为信号序列的额外受体。我们测量了N(ub)标记的Ssh1p与带有四种不同信号序列的C(ub)易位底物之间的相互作用。将如此确定的Ssh1p的相互作用谱与相应修饰的N(ub)-Sec61p的信号序列相互作用谱进行比较。该测定揭示了Ssh1p与Kar2p和蔗糖酶的信号序列之间的相互作用，而Sec61p还与Mfalpha1和羧肽酶Y的信号序列相互作用。所测量的Ssh1p与信号序列识别颗粒受体的β亚基之间的物理接近性证实了我们的假设，即Ssh1p直接参与蛋白质在内质网膜上的共翻译易位。
