Popoff M R, Chaves-Olarte E, Lemichez E, von Eichel-Streiber C, Thelestam M, Chardin P, Cussac D, Antonny B, Chavrier P, Flatau G, Giry M, de Gunzburg J, Boquet P
Institut Pasteur, Unité des Toxines Microbiennes, 75724 Paris, Cedex 15, France.
J Biol Chem. 1996 Apr 26;271(17):10217-24. doi: 10.1074/jbc.271.17.10217.
Lethal toxin (LT) from Clostridium sordellii is one of the high molecular mass clostridial cytotoxins. On cultured cells, it causes a rounding of cell bodies and a disruption of actin stress fibers. We demonstrate that LT is a glucosyltransferase that uses UDP-Glc as a cofactor to covalently modify 21-kDa proteins both in vitro and in vivo. LT glucosylates Ras, Rap, and Rac. In Ras, threonine at position 35 was identified as the target amino acid glucosylated by LT. Other related members of the Ras GTPase superfamily, including RhoA, Cdc42, and Rab6, were not modified by LT. Incubation of serum-starved Swiss 3T3 cells with LT prevents the epidermal growth factor-induced phosphorylation of mitogen-activated protein kinases ERK1 and ERK2, indicating that the toxin blocks Ras function in vivo. We also demonstrate that LT acts inside the cell and that the glucosylation reaction is required to observe its dramatic effect on cell morphology. LT is thus a powerful tool to inhibit Ras function in vivo.
来自索氏梭菌的致死毒素（LT）是高分子量梭菌细胞毒素之一。在培养细胞上，它会导致细胞体变圆并破坏肌动蛋白应力纤维。我们证明LT是一种葡糖基转移酶，它以UDP - 葡萄糖作为辅因子，在体外和体内共价修饰21 kDa的蛋白质。LT使Ras、Rap和Rac葡糖基化。在Ras中，第35位的苏氨酸被确定为被LT葡糖基化的目标氨基酸。Ras GTPase超家族的其他相关成员，包括RhoA、Cdc42和Rab6，未被LT修饰。用LT孵育血清饥饿的瑞士3T3细胞可阻止表皮生长因子诱导的丝裂原活化蛋白激酶ERK1和ERK2的磷酸化，表明该毒素在体内阻断Ras功能。我们还证明LT在细胞内起作用，并且葡糖基化反应是观察其对细胞形态产生显著影响所必需的。因此，LT是一种在体内抑制Ras功能的强大工具。
