Translocation of a long amino-terminal domain through ER membrane by following signal-anchor sequence
Translocation of a long amino-terminal domain through ER membrane by following signal-anchor sequence
复制标题
DOI：
10.1038/sj.emboj.7600788
复制
发表时间：
2005-09-21
期刊：
影响因子：
11.4
通讯作者：
Sakaguchi, M
中科院分区：
文献类型：
--
作者：
Kida, Y;Mihara, K;Sakaguchi, M
I类信号锚序列介导N-末端结构域(N-结构域)跨内质网(ER)膜的移位。为了详细研究移位，二氢叶酸还原酶(DHFR)被融合到突触素II的N-末端作为一个长的N-结构域。转位被DHFR配体甲氨蝶呤阻止，它稳定了DHFR结构域的折叠，并在甲氨蝶呤耗尽后恢复。核糖体-新生链状复合体靶向内质网需要GTP，而N-结构域的易位不需要任何核苷酸三磷酸。即使在没有管腔热休克蛋白70(BiP)的情况下也观察到了显著的易位。当膜靶向后从核糖体中释放新生多肽时，N-结构域的易位被抑制，新生的链从易位子中释放出来。核糖体在维持易位中间状态方面起着至关重要的作用。当dhfr结构域从信号锚序列中分离出来时，它的易位受到了极大的损害。DHFR结构域的展开和移位必须由信号锚序列进入易位子来驱动。
Type I signal-anchor sequences mediate translocation of the N-terminal domain (N-domain) across the endoplasmic reticulum ( ER) membrane. To examine the translocation in detail, dihydrofolate reductase ( DHFR) was fused to the N-terminus of synaptotagmin II as a long N-domain. Translocation was arrested by the DHFR ligand methotrexate, which stabilizes the folding of the DHFR domain, and resumed after depletion of methotrexate. The targeting of the ribosome - nascent chain complex to the ER requires GTP, whereas N-domain translocation does not require any nucleotide triphosphates. Significant translocation was observed even in the absence of a lumenal hsp70 (BiP). When the nascent polypeptide was released from the ribosomes after the membrane targeting, the N-domain translocation was suppressed and the nascent chain was released from the translocon. Ribosomes have a crucial role in maintaining the translocation-intermediate state. The translocation of the DHFR domain was greatly impaired when it was separated from the signal-anchor sequence. Unfolding and translocation of the DHFR domain must be driven by the stroke of the signal-anchor sequence into translocon.