Membrane cholesterol and sphingomyelin, and ostreolysin A are obligatory for pore-formation by a MACPF/CDC-like pore-forming protein, pleurotolysin B

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• 作者：Katja Ota ; Adrijana Leonardi ; Miha Mikelj ; Matej Sko?aj ; Therese Wohlschlager ; Markus K¨¹nzler ; Markus Aebi ; Mojca Narat ; Igor Kri?aj ; Gregor Anderluh ; Kristina Sep?i? ; Peter Ma?ek
• 关键词：Aegerolysin ; Cholesterol-dependent cytolysin ; Lipid-binding protein ; Membrane attack complex ; Pleurotolysin ; Pleurotus ostreatus
• 刊名：Biochimie
• 出版年：2013
• 出版时间：October, 2013
• 年：2013
• 卷：95
• 期：10
• 页码：1855-1864
• 全文大小：1672 K

文摘

The mushroom Pleurotus ostreatus has been reported to produce the hemolytic proteins ostreolysin (OlyA), pleurotolysin A (PlyA) and pleurotolysin B (PlyB). The present study of the native and recombinant proteins dissects out their lipid-binding characteristics and their roles in lipid binding and membrane permeabilization. Using lipid-binding studies, permeabilization of erythrocytes, large unilamellar vesicles of various lipid compositions, and electron microscopy, we show that OlyA, a PlyA homolog, preferentially binds to membranes rich in sterol and sphingomyelin, but it does not permeabilize them. The N-terminally truncated ¦¤48PlyB corresponds to the mature and active form of native PlyB, and it has a membrane attack complex-perforin (MACPF) domain. ¦¤48PlyB spontaneously oligomerizes in solution, and binds weakly to various lipid membranes but is not able to perforate them. However, binding of ¦¤48PlyB to the cholesterol and sphingomyelin membranes, and consequently, their permeabilization is dramatically promoted in the presence of OlyA. On these membranes, ¦¤48PlyB and OlyA form predominantly 13-meric oligomers. These are rosette-like structures with a thickness of ¡«9?nm from the membrane surface, with 19.7?nm and 4.9?nm outer and inner diameters, respectively. When present on opposing vesicle membranes, these oligomers can dimerize and thus promote aggregation of vesicles. Based on the structural and functional characteristics of ¦¤48PlyB, we suggest that it shares some features with MACPF/cholesterol-dependent cytolysin (CDC) proteins. OlyA is obligatory for the ¦¤48PlyB permeabilization of membranes rich in cholesterol and sphingomyelin.