Structural and biophysical insights into the role of the insert region in Rac1 function

摘要: A 13 amino acid insertion that forms a short 3(10) helix between beta-strand 5 and alpha-helix 4 is distinguishing feature among most members of the Rho family GTPases, yet precise role this region in signal transduction poorly understood. Previous vivo functional studies have implicated insert RhoA, Rac1, Cdc42 to be important for cell transformation, regulation actin cytoskeleton, controlling DNA synthesis, activation downstream targets. In our recent biological suggested SRF formation lamellipodia but dispensable all other cellular functions protein. reported herein, we characterized effect deletion on Rac1 structure, thermodynamic stability, kinetics nucleotide association. These vitro help clarify data provide further insights as modulating function. The reveal has no structure causes only marginal (approximately 0.8 kcal/mol) decrease DeltaG(fold) Rac1*GDP*Mg2+. intrinsic rate dissociation Rac1*Delta(ins) decreased by about 1.5-fold compared wild type, 3-fold increase GEF (Vav2)-mediated exchange observed. addition, does not change K(D) interaction with GDI, similar previously observed Cdc42, inhibition GDP mutant relative native Taken together, structural biochemical here are consistent suggest likely must serve binding interface effectors, particularly those regulation.