Simon S M, Blobel G
Laboratory of Cell Biology, Howard Hughes Medical Institute, Rockefeller University, New York, New York 10021-6399.
Cell. 1991 May 3;65(3):371-80. doi: 10.1016/0092-8674(91)90455-8.
The existence of a protein-conducting channel in the endoplasmic reticulum membrane was demonstrated by electrophysiological techniques. Pancreatic rough microsome (RM) vesicles were fused to one side (cis) of a planar lipid bilayer separating two aqueous compartments of 50 mM salt. This exposed the cytoplasmic surface of the RMs, with its attached ribosomes, to the cis chamber. Addition of 100 microM puromycin to the cis side caused a large increase in membrane conductance, presumably the result of puromycin-induced clearance of nascent protein chains from the lumen of protein-conducting channels. When puromycin was added at low concentrations (0.33 microM), single channels of 220 pS were observed. These closed when the salt concentration was raised to levels at which ribosomes detach from the membrane (150-400 mM), indicating that the attached ribosome keeps the channel in an open conformation. A mechanism for a complete cycle of opening and closing of the protein-conducting channel is suggested.
通过电生理技术证实了内质网膜中存在蛋白质传导通道。将胰腺粗微粒体（RM）囊泡融合到分隔两个50 mM盐溶液水相区室的平面脂质双层的一侧（顺式）。这使得带有附着核糖体的RM的细胞质表面暴露于顺式腔室。向顺式侧添加100 microM嘌呤霉素会导致膜电导大幅增加，这可能是嘌呤霉素诱导新生蛋白质链从蛋白质传导通道腔中清除的结果。当以低浓度（0.33 microM）添加嘌呤霉素时，观察到了220 pS的单通道。当盐浓度升高到核糖体从膜上脱离的水平（150 - 400 mM）时，这些通道关闭，表明附着的核糖体使通道保持开放构象。提出了蛋白质传导通道完整开闭循环的机制。
