University of Osnabrück, Barbarastrasse 7, D-49069 Osnabrück, Germany.
J Mol Biol. 2013 May 27;425(10):1782-94. doi: 10.1016/j.jmb.2013.01.037. Epub 2013 Feb 8.
Colicin A is a pore-forming toxin that forms a voltage-gated channel in the inner membrane of the target bacteria. The structures of the closed and open channel states of membrane-bound colicin A are not resolved. In the present site-directed spin-labeling study, the insertion-competent state of colicin A is provoked by an acidic pH jump prior to the insertion into liposomes prepared from Escherichia coli natural lipids. The membrane-bound colicin A is able to open a voltage-dependent channel as demonstrated by the efflux of tempophosphate spin label from the lumen of liposomes. The EPR spectra of spin-labeled colicin A variants in the membrane-bound closed channel state reveal a conformational equilibrium with resolved interhelical tertiary contacts. The spin label accessibility and polarity profiles suggest the amphipathic helices (H1-H7 and H10) to be located in the membrane close to the membrane-water interface and the hydrophobic hairpin (H8 and H9) to be immersed more deeply in the membrane.
大肠杆菌素 A 是一种形成孔的毒素，它在靶细菌的内膜中形成一个电压门控通道。结合膜的大肠杆菌素 A 的关闭和开放通道状态的结构尚未解决。在本定点自旋标记研究中，在插入脂质体之前，通过酸性 pH 跳跃引发插入能力状态的大肠杆菌素 A，该脂质体由大肠杆菌天然脂质制成。膜结合的大肠杆菌素 A 能够打开电压依赖性通道，这通过从脂质体腔中 tempophosphate 自旋标记的流出来证明。在膜结合的关闭通道状态下的突变体的 EPR 光谱揭示了具有分辨的螺旋间三级接触的构象平衡。自旋标记的大肠杆菌素 A 变体的自旋标记可及性和极性分布表明两亲性螺旋（H1-H7 和 H10）位于靠近膜 - 水界面的膜中，疏水性发夹（H8 和 H9）更深入地浸入膜中。
