BIOSYNTHESIS AND FUNCTION OF GLYCOSPHINGOLIPIDS AND OTHER GLYCOCONJUGATES

神经节苷脂G（M1）是霍乱毒素（CT）的细胞表面受体。 寡糖链（G（M1）OS）被B或结合 CT的A亚单位，而毒素的A亚单位激活腺苷酸 通过ADP-核糖基化的刺激性G蛋白（Gs）的环化酶 环化酶系统 A亚基进入G的途径 尚未建立。 我们之前已经证明， 通过将G（M1）OS共价连接至细胞产生的新神经节蛋白 表面蛋白是CT的无功能受体。 因此，CT结合大鼠 用G（M1）OS修饰但不能刺激腺苷酸的胶质瘤C6 环化酶 我们最近发现，在氯喹的存在下，这些 新神经节蛋白成为CT的功能性受体。 进一步 为了探索新神经节细胞蛋白作为毒素受体的作用，我们通过共价键 将G（M1）OS连接至人转铁蛋白。 导数与高 与HeLa细胞上的转铁蛋白受体的亲和力，并支持结合 的CT。 然而，毒素不能激活腺苷酸环化酶， 与G（M1）处理的HeLa细胞相比。 氯喹治疗允许CTL 在用G（M1）-转铁蛋白处理的HeLa细胞中激活腺苷酸环化酶。 相比之下，对照或G（M1）处理的HeLa或C6细胞的预处理， 氯喹对毒素刺激腺苷酸的作用无影响 环化酶 这些结果表明，与G（M1）结合的CT获得了对G（M1）的访问。 腺苷酸环化酶通过不同的途径，而不是结合时， 新神经节蛋白 E.大肠杆菌不耐热肠毒素（LT）的结构是 类似于CT，结合GM，并激活腺苷酸环化酶。 人类 肠细胞系被发现具有额外的LT结合位点， 刷状缘膜。 这些地点被确定为 半乳糖蛋白 我们现在正在确定这些结合位点是否可以 作为LT的功能性受体并允许LT进入细胞 通过与CT不同的途径

Ganglioside G(M1) is the cell surface receptor for cholera toxin (CT). The oligosaccharide chain (G(M1)OS) is recognized by the B or binding subunits of CT whereas the A subunit of the toxin activates adenylyl cyclase by ADP-ribosylation of the stimulatory G protein (Gs) of the cyclase system. The pathway by which the A subunit gains access to Gs has not yet been established. We previously had shown that neoganglioproteins generated by covalently attaching G(M1)OS to cell surface proteins are nonfunction receptors for CT. Thus, CT binds to rat glioma C6 modified with G(M1)OS but is unable to stimulate adenylyl cyclase. We recently found that in the presence of chloroquine, these neoganglioproteins became functional receptors for CT. To further explore the role of neoganglioproteins as toxin receptors,we covalently attached G(M1)OS to human transferrin. The derivative bound with high affinity to transferrin receptors on HeLa cells and supported the binding of CT. The toxin, however, was unable to activate adenylyl cyclase in contrast to G(M1)-treated HeLa cells. Chloroquine treatment allowed CTL activate adenylyl cyclase in HeLa cells treated with G(M1)-transferrin. By contrast, pretreatment of either control or G(M1)-treated HeLa or C6 cells with chloroquine had no effect on toxin stimulation of adenylyl cyclase. These results indicate that CT bound to G(M1) gains access to adenylyl cyclase through a different pathway than when bound to neoganglioproteins. E. coli heat-labile enterotoxin (LT) is structurally similar to CT, binds to GM, and activates adenylyl cyclase. A human intestinal cell line was found to have additional binding sites for LT on the brush border membranes. These sites were identified as galactoproteins. We are now determining whether these binding sites can serve as functional receptors for LT and allow LT to enter the cells through a different pathway than CT.