Chemical and photochemical modification of colicin E1 and gramicidin A in bilayer lipid membranes.
Chemical and photochemical modification of colicin E1 and gramicidin A in bilayer lipid membranes.
复制标题
双层脂膜中大肠菌素 E1 和短杆菌肽 A 的化学和光化学修饰。
DOI：
10.1007/s00232-004-0674-y
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Antonenko,YN
中科院分区：
文献类型：
--
作者：
Sobko,AA;Vigasina,MA;Rokitskaya,TI;Kotova,EA;Zakharov,SD;Cramer,WA;Antonenko,YN
采用化学修饰和光动力处理的方法研究了水疱和平面双层脂质膜（BLMs）中colicin E1通道形成结构域（P178），以阐明色氨酸残基在colicin E1通道活性中的作用。用n -溴琥珀酰亚胺（NBS）修饰大肠杆菌色氨酸残基，从色氨酸荧光的丧失判断，在由完全饱和（二植烷酰）磷脂形成的BLMs中，在宏观电流和单通道水平上，导致野生型P178通道活性完全抑制。NBS处理革兰杀菌素通道后，对BLM上的色氨酸荧光和电流也有类似的影响。在所研究的单色氨酸P178突变体中，W460对NBS处理表现出最高的敏感性，这表明暴露于水的Trp460对大肠杆菌素通道活性的重要性。与之前的研究一致，光动力处理（在光敏剂存在下的可见光照射）导致双植烷酰磷脂膜中P178通道活性的抑制，同时通过色氨酸荧光的减少检测到色氨酸残基的损伤。目前的工作揭示了新的效应：由不饱和（二油基）磷脂形成的blm可以观察到NBS和光动力处理对P178通道的激活。这些现象归因于双键脂质氧化修饰对P178通道形成的影响。在NBS预处理和BLMs敏化后，观察到大肠杆菌素介导的离子电流明显受到刺激，这支持了膜结构扭曲可以促进通道形成的观点。
Chemical modification and photodynamic treatment of the colicin E1 channel-forming domain (P178) in vesicular and planar bilayer lipid membranes (BLMs) was used to elucidate the role of tryptophan residues in colicin E1 channel activity. Modification of colicin tryptophan residues by N-bromosuccinimide (NBS), as judged by the loss of tryptophan fluorescence, resulted in complete suppression of wild-type P178 channel activity in BLMs formed from fully saturated (diphytanoyl) phospholipids, both at the macroscopic-current and single-channel levels. The similar effect on both the tryptophan fluorescence and the electric current across BLM was observed also after NBS treatment of gramicidin channels. Of the single-tryptophan P178 mutants studied, W460 showed the highest sensitivity to NBS treatment, pointing to the importance of the water-exposed Trp460 in colicin channel activity. In line with previous work, the photodynamic treatment (illumination with visible light in the presence of a photosensitizer) led to suppression of P178 channel activity in diphytanoyl-phospholipid membranes concomitant with the damage to tryptophan residues detected here by a decrease in tryptophan fluorescence. The present work revealed novel effects: activation of P178 channels as a result of both NBS and photodynamic treatments was observed with BLMs formed from unsaturated (dioleoyl) phospholipids. These phenomena are ascribed to the effect of oxidative modification of double-bond-containing lipids on P178 channel formation. The pronounced stimulation of the colicin-mediated ionic current observed after both pretreatment with NBS and sensitized photomodification of the BLMs support the idea that distortion of membrane structure can facilitate channel formation.