van den Berghe N, Cool R H, Wittinghofer A
Max-Planck-Institut für Molekulare Physiologie, Abteilung Strukturelle Biologie, Rheinlanddamm 201, 44139 Dortmund, Germany.
J Biol Chem. 1999 Apr 16;274(16):11078-85. doi: 10.1074/jbc.274.16.11078.
The inability of the S17N mutant of Rap1A to sequester the catalytic domain of the Rap guanine nucleotide exchange factor C3G (van den Berghe, N., Cool, R. H., Horn, G., and Wittinghofer, A. (1997) Oncogene 15, 845-850) prompted us to study possible fundamental differences in the way Rap1 interacts with C3G compared with the interaction of Ras with the catalytic domain of the mouse Ras guanine nucleotide exchange factor Cdc25(Mm). A variety of mutants in both Ras and Rap1A were designed, and both the C3G and Cdc25(Mm) catalyzed release of guanine nucleotide from these mutants was studied. In addition, we could identify regions in Rap2A that are responsible for the lack of recognition by C3G and induce high C3G activity by replacement of these residues with the corresponding Rap1A residues. The different Ras and Rap mutants showed that many residues were equally important for both C3G and Cdc25(Mm), suggesting that they interact similarly with their substrates. However, several residues were also identified to be important for the exchange reaction with only C3G (Leu70) or only Cdc25(Mm) (Gln61 and Tyr40). These results are discussed in the light of the structure of the Ras-Sos complex and suggest that some important differences in the interaction of Rap1 with C3G and Ras with Cdc25(Mm) indeed exist and that marker residues have been identified for the different structural requirements.
Rap1A的S17N突变体无法隔离Rap鸟嘌呤核苷酸交换因子C3G的催化结构域（范登伯格，N.，库尔，R. H.，霍恩，G.，和维廷霍费尔，A.（1997年）《癌基因》15卷，845 - 850页），这促使我们研究Rap1与C3G相互作用的方式与Ras与小鼠Ras鸟嘌呤核苷酸交换因子Cdc25（Mm）的催化结构域相互作用方式之间可能存在的根本差异。我们设计了Ras和Rap1A中的多种突变体，并研究了C3G和Cdc25（Mm）催化这些突变体释放鸟嘌呤核苷酸的情况。此外，我们能够确定Rap2A中负责不被C3G识别的区域，并通过用相应的Rap1A残基替换这些残基来诱导高C3G活性。不同的Ras和Rap突变体表明，许多残基对C3G和Cdc25（Mm）同样重要，这表明它们与底物的相互作用方式相似。然而，也确定了几个仅对与C3G的交换反应（Leu70）或仅对与Cdc25（Mm）的交换反应（Gln61和Tyr40）重要的残基。根据Ras - Sos复合物的结构对这些结果进行了讨论，结果表明Rap1与C3G的相互作用和Ras与Cdc25（Mm）的相互作用确实存在一些重要差异，并且已经确定了针对不同结构要求的标记残基。
