Cdc42 / Cdc42GAP complex
Homo sapiens (human)
G-proteins (GTP-binding proteins, guanine nucleotide binding proteins) and GTPase-activating proteins (GAPs) are essential component of all signaling pathways mediated by Ras superfamily. G-proteins act as molecular switches, which is active in GTP-bound state and inactive in GDP-bound state. The GTPase activity of G-proteins allows them to cycle between the active state and the inactive state. Although the GTPase activity of G-proteins is generally low, GAPs can enhance the hydrolysis rate by up to hundred thousand times, whose mechanism is not known well. Aluminum fluoride (AlF4-) is known to have an ability to stimulate activation of G-protein. Upon binding of AlF4- to inactive state G-proteins, G-protein adopts the conformation of active state, which act as a transition state analog and enables signaling to downstream effector. The authors of this paper previously found that aluminum fluoride can induce stable complex of the Cdc42 and Cdc42GAP, but this transition state-like structure haven't been solved.
The structure shown here is the complex of Cdc42 and Cdc42GAP. Cdc42 adopts the classical G-domain fold. The interaction between Cdc42 and Cdc42GAP is mediated by the region called switch I and switch II in Cdc42 and residue300-310 in Cdc42GAP, so-called finger loop. C-terminal region forms a long solvent-exposed loop, which is stabilized by disulfide bridge between Cys105 and Cys188 and by the hydrogen-bond interactions including contacts with residues in switch II. Aluminum fluoride is observed in active site of the Cdc42, mimicking the penta-covalent transition state structure in the GTP-hydrolyric pathway and making hydrogen bond with catalytic arginine (Arg305) in Cdc42GAP. The authors of this paper induced mutation in Arg305 to Ala and found that the loss of Arg305 leads to an increased mobility of finger loop and weakening or loss of hydrogen bonds between finger loop and the switch II region, result in reduced affinity to Cdc42. They indicated that GAP introduces the catalytically critical arginine, which serves to stabilize transition state and excluding solvent from active site to maintain catalysis of GTP hydrolysis.
Protein Data Bank (PDB)
author: Miho Higurashi