A PROTEIN-CONDUCTING CHANNEL IN THE ENDOPLASMIC-RETICULUM

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-mu-M 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-mu-M), 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.