Ishitsuka Reiko, Kobayashi Toshihide
Lipid Biology Laboratory, RIKEN (Institute of Physical and Chemical Research) Discovery Research Institute, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan.
Biochemistry. 2007 Feb 13;46(6):1495-502. doi: 10.1021/bi061290k. Epub 2007 Jan 23.
Lysenin is a pore-forming toxin that specifically binds sphingomyelin (SM). The binding of the toxin to the membrane is accompanied by the oligomerization of the protein, leading to pore formation. The interaction of lysenin with SM is affected by the presence of other lipids found in the plasma membrane. Although a previous study showed that SM/cholesterol liposomes were 10,000 times more effective than SM liposomes in inhibiting lysenin-induced hemolysis (Yamaji, A., Sekizawa, Y., Emoto, K., Sakuraba, H., Inoue, K., Kobayashi, H., and Umeda, M. (1998) J. Biol. Chem. 273, 5300-5306), the role of cholesterol is not precisely clarified. In the present study, we examined the effects of the presence of cholesterol in the SM membrane on the inhibition of hemolysis, the binding of lysenin to SM, and the oligomerization of lysenin. The addition of cholesterol to SM liposomes dramatically inhibited lysenin-induced hemolysis as described previously. However, the presence of cholesterol did not affect the binding of lysenin to SM liposomes. The oligomerization of lysenin was facilitated by the presence of cholesterol in SM liposomes. The oligomerization of lysenin was also dependent on the SM/lysenin ratio, that is, the amount of lysenin oligomer was increased with the decrease in the SM/lysenin ratio. When the SM/lysenin molar ratio was high, lysenin associated with the membrane as a monomer, which was able to transfer to the erythrocyte membrane. Our results indicate that both cholesterol and the SM/lysenin ratio control the amount of lysenin monomer via altering the state of protein oligomerization, thus affecting hemolysis.
溶细胞素是一种能特异性结合鞘磷脂（SM）的成孔毒素。毒素与膜的结合伴随着蛋白质的寡聚化，导致孔的形成。溶细胞素与SM的相互作用受质膜中其他脂质存在的影响。尽管先前的一项研究表明，SM/胆固醇脂质体在抑制溶细胞素诱导的溶血方面比SM脂质体有效10000倍（山治，A.，关泽，Y.，江本，K.，樱叶，H.，井上，K.，小林，H.，以及梅田，M.（1998年）《生物化学杂志》273卷，5300 - 5306页），但胆固醇的作用尚未得到确切阐明。在本研究中，我们研究了SM膜中胆固醇的存在对溶血抑制、溶细胞素与SM的结合以及溶细胞素寡聚化的影响。如前所述，向SM脂质体中添加胆固醇可显著抑制溶细胞素诱导的溶血。然而，胆固醇的存在并不影响溶细胞素与SM脂质体的结合。SM脂质体中胆固醇的存在促进了溶细胞素的寡聚化。溶细胞素的寡聚化也取决于SM/溶细胞素的比例，即溶细胞素寡聚体的量随着SM/溶细胞素比例的降低而增加。当SM/溶细胞素摩尔比高时，溶细胞素以单体形式与膜结合，该单体能够转移到红细胞膜上。我们的结果表明，胆固醇和SM/溶细胞素比例均通过改变蛋白质寡聚化状态来控制溶细胞素单体的量，从而影响溶血。
