Abraham Sherwin J, Nolet Ryan P, Calvert Richard J, Anderson Lucy M, Gaponenko Vadim
Department of Biochemistry and Molecular Genetics, University of Illinois, Chicago, Illinois 60607, USA.
Biochemistry. 2009 Aug 18;48(32):7575-83. doi: 10.1021/bi900769j.
K-Ras4B belongs to the family of p21 Ras GTPases, which play an important role in cell proliferation, survival, and motility. The p21 Ras proteins, such as K-Ras4B, K-Ras4A, H-Ras, and N-Ras, share 85% sequence homology and activate very similar signaling pathways. Only the C-terminal hypervariable regions differ significantly. A growing body of literature demonstrates that each Ras isoform possesses unique functions in normal physiological processes as well as in pathogenesis. One of the central questions in the field of Ras biology is how these very similar proteins achieve such remarkable specificity in protein-protein interactions that regulate signal transduction pathways. Here we explore specific binding of K-Ras4B to calmodulin. Using NMR techniques and isothermal titration calorimetry, we demonstrate that the hypervariable region of K-Ras4B contributes in a major way to the interaction with calmodulin, while the catalytic domain of K-Ras4B provides a way to control the interaction by nucleotide binding. The hypervariable region of K-Ras4B binds specifically to the C-terminal domain of Ca(2+)-loaded calmodulin with micromolar affinity, while the GTP-gamma-S-loaded catalytic domain of K-Ras4B may interact with the N-terminal domain of calmodulin.
K-Ras4B属于p21 Ras GTP酶家族，该家族在细胞增殖、存活和运动中起重要作用。p21 Ras蛋白，如K-Ras4B、K-Ras4A、H-Ras和N-Ras，具有85%的序列同源性，并激活非常相似的信号通路。只有C末端高变区有显著差异。越来越多的文献表明，每种Ras异构体在正常生理过程以及发病机制中都具有独特的功能。Ras生物学领域的核心问题之一是，这些非常相似的蛋白质如何在调节信号转导通路的蛋白质-蛋白质相互作用中实现如此显著的特异性。在这里，我们探索K-Ras4B与钙调蛋白的特异性结合。使用核磁共振技术和等温滴定量热法，我们证明K-Ras4B的高变区在很大程度上有助于与钙调蛋白的相互作用，而K-Ras4B的催化结构域通过核苷酸结合提供了一种控制相互作用的方式。K-Ras4B的高变区以微摩尔亲和力特异性结合Ca(2+)负载的钙调蛋白的C末端结构域，而K-Ras4B的GTP-γ-S负载的催化结构域可能与钙调蛋白的N末端结构域相互作用。
