Glucosylation and ADP ribosylation of Rho proteins: Effects on nucleotide binding, GTPase activity, and effector coupling
Glucosylation and ADP ribosylation of Rho proteins: Effects on nucleotide binding, GTPase activity, and effector coupling
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DOI：
10.1021/bi972592c
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发表时间：
1998-04-14
期刊：
影响因子：
2.9
通讯作者：
Aktories, K
中科院分区：
文献类型：
--
作者：
Sehr, P;Joseph, G;Aktories, K
我们研究了艰难梭菌毒素 B 对 RhoA、Rac1 和 Cdc42 在苏氨酸 35 和 -37 处的糖基化对核苷酸结合、GTP 酶活性和效应子偶联的影响，并将这些结果与肉毒杆菌 C3 转移酶催化的 RhoA 在天冬酰胺 41 处的 ADP 核糖基化进行了比较。而葡萄糖基化和 ADP 核糖基化对 RhoA、Rac1 和 Cdc42 的 GDP 释放没有重大影响。通过糖基化，Rho 蛋白释放 GTP gamma S 的速率增加了 3-6 倍。 ADP 核糖基化使 GTP gamma S 释放率降低约 50%。糖基化使 GTPase 的内在活性降低 3-7 倍，并完全阻断 RhoGAP 的 GTPase 刺激。相反，ADP 核糖基化略微增加了 GTPase 活性（接近 2 倍），并且对 GTPase 的 GAP 刺激没有重大影响。尽管 ADP 核糖基化不影响 RhoA 与蛋白激酶 N 结合域的相互作用，但糖基化却抑制了这种相互作用。 Rac1 的糖基化显着降低了其支持吞噬细胞产生超氧化物的 NADPH 氧化酶激活的能力。糖基化的 Rac1 不会干扰未修饰的 Rac1 的 NADPH 氧化酶激活，即使存在明显摩尔过量时，这表明它无法竞争共同的效应子。数据表明，糖基化引起的小 GTP 酶功能失活主要是由 GTP 酶-效应蛋白相互作用的抑制引起的。
We studied the effects of glucosylation of RhoA, Rac1, and Cdc42 at threonine-35 and -37 by Clostridium difficile toxin B on nucleotide binding, GTPase activity, and effector coupling and compared these results with the ADP ribosylation of RhoA at asparagine-41 catalyzed by Clostridium botulinum C3 transferase. Whereas glucosylation and ADP ribosylation had no major effects on GDP release from RhoA, Rac1, and Cdc42. the rate of GTP gamma S release from Rho proteins was increased 3-6-fold by glucosylation. ADP ribosylation decreased the rate of GTP gamma S release by about 50%. Glucosylation reduced the intrinsic activities of the GTPases by 3-7-fold and completely blocked GTPase stimulation by RhoGAP. In contrast, ADP ribosylation slightly increased GTPase activity (similar to 2-fold) and had no major effect on GAP stimulation of GTPase. Whereas ADP ribosylation did not affect the interaction of RhoA with the binding domain of protein kinase N, glucosylation inhibited this interaction. Glucosylation of Rac1 markedly diminished its ability to support the activation of the superoxide-generating NADPH oxidase of phagocytes. Glucosylated Rac1 did not interfere with NADPH oxidase activation by unmodified Rac1, even when present in marked molar excess, indicating that it was incapable of competing for a common effector. The data indicate that the functional inactivation of small GTPases by glucosylation is mainly caused by inhibition of GTPase-effector protein interaction.