Slatin S L, Raymond L, Finkelstein A
J Membr Biol. 1986;92(3):247-54. doi: 10.1007/BF01869393.
The voltage-dependent channel formed in planar lipid bilayers by colicin E1, or its channel-forming C-terminal fragments, is susceptible to destruction by the nonspecific protease pepsin under well-defined conditions. In particular, pepsin acts only from the cis side (the side to which colicin has been added) and only upon channels in the closed state. Channels in the open state are refractory to destruction by cis pepsin, and neither open nor closed channels are destroyed by trans pepsin. Colicin E1 channels are normally turned on by cis positive voltages and turned off by cis negative voltages. For large (greater than 80 mV) positive voltages, however, channels inactivate subsequent to opening. Associated with the inactivated state, some channels become capable of being turned on by cis negative voltages and turned off by cis positive voltages, as if the channel-forming region of the molecule has been translocated across the membrane. Consistent with this interpretation is the ability now of trans pepsin to destroy these "reversed" channels when they are closed, but not when they are open, whereas cis pepsin has no effect on them in either the open or closed state. Our results indicate that voltage gating of the E1 channel involves translocation of parts of the protein across the membrane, exposing different domains to the cis and trans solutions in the different channel states.
由大肠杆菌素E1或其形成通道的C末端片段在平面脂质双分子层中形成的电压依赖性通道，在明确的条件下易受非特异性蛋白酶胃蛋白酶的破坏。具体而言，胃蛋白酶仅从顺式侧（添加大肠杆菌素的一侧）起作用，并且仅作用于处于关闭状态的通道。处于开放状态的通道对顺式胃蛋白酶的破坏具有抗性，并且无论是开放通道还是关闭通道都不会被反式胃蛋白酶破坏。大肠杆菌素E1通道通常由顺式正电压开启，由顺式负电压关闭。然而，对于大的（大于80 mV）正电压，通道在开放后会失活。与失活状态相关，一些通道变得能够被顺式负电压开启并被顺式正电压关闭，就好像分子的通道形成区域已经跨膜移位。与此解释一致的是，当这些“反向”通道关闭时，反式胃蛋白酶能够破坏它们，但在开放时则不能，而顺式胃蛋白酶在开放或关闭状态下对它们都没有影响。我们的结果表明，E1通道的电压门控涉及蛋白质部分跨膜移位，在不同的通道状态下将不同的结构域暴露于顺式和反式溶液中。
