Prepore to pore transition of a cholesterol-dependent cytolysin visualized by electron microscopy
Prepore to pore transition of a cholesterol-dependent cytolysin visualized by electron microscopy
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DOI：
10.1016/j.jsb.2005.02.003
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发表时间：
2005-04-01
影响因子：
3
通讯作者：
Wilson-Kubalek, EM
中科院分区：
文献类型：
--
作者：
Dang, TX;Hotze, EM;Wilson-Kubalek, EM
产气链球菌溶血素O（PFO）是一种由致病性培氏梭菌分泌的可溶性毒素，在含胆固醇的膜中形成由多达50个PFO分子组成的大型同源寡聚孔复合物。在这项研究中，电子显微镜（EM）和单粒子图像分析被用来重建二维（213）投影图从低聚PFO前孔和孔复合物上形成的富含胆固醇的脂质层的图像。投影图的特征在于密度峰值的外环和内环。前孔复合物和孔复合物的外环非常相似;然而，构成孔复合物内环的蛋白质密度比前孔复合物的蛋白质密度更强且分辨率更离散。内环蛋白质密度的变化与其中前孔复合物内的单体在孔复合物中从部分无序状态转变为更有序的跨膜β-桶的机制一致。最后，寡聚复合物内的单体的取向通过可视化链霉亲和素（SA）分子结合到生物素化的半胱氨酸取代的残基预测面对寡聚孔复合物的内表面或外表面来确定。这项研究提供了一个前所未有的观点的PFO前孔的转换孔复合物。(c)2005年爱思唯尔公司All rights reserved.
Perfringolysin O (PFO), a soluble toxin secreted by the pathogenic Clostridium peifringens, forms large homo-oligomeric pore complexes comprising up to 50 PFO molecules in cholesterol-containing membranes. In this study, electron microscopy (EM) and single-particle image analysis were used to reconstruct two-dimensional (213) projection maps from images of oligomeric PFO pre-pore and pore complexes formed on cholesterol-rich lipid layers. The projection maps are characterized by an outer and an inner ring of density peaks. The outer rings of the prepore and pore complexes are very similar; however, the protein densities that make up the inner ring of the pore complex are more intense and discretely resolved than they are for the prepore complex. The change in inner-ring protein density is consistent with a mechanism in which the monomers within the prepore complex make a transition from a partially disordered state to a more ordered transmembrane beta-barrel in the pore complex. Finally, the orientation of the monomers within the oligomeric complexes was determined by visualization of streptavidin (SA) molecules bound to biotinylated cysteine-substituted residues predicted to face either the inner or outer surface of the oligomeric pore complex. This study provides an unprecedented view of the conversion of the PFO prepore to pore complex. (c) 2005 Elsevier Inc. All rights reserved.