Members of the Rho family of small G proteins transduce signals from plasma-membrane receptors and control cell adhesion, motility and shape by actin cytoskeleton formation(1-4), They also activate other kinase cascades, Like all other GTPases, Rho proteins act as molecular switches, with an active GTP-bound form and an inactive GDP-bound form(5), The active conformation is promoted by guanine-nucleotide exchange factors, and the inactive state by GTPase-activating proteins (GAPs) which stimulate the intrinsic GTPase activity of small G proteins(6), Rho-specific GAP domains are found in a wide variety of large, multi-functional proteins(7), Here we report the crystal structure of an active 242-residue C-terminal fragment of human p50rhoGAP(8), The structure is an unusual arrangement of nine alpha-helices, the core of which includes a four-helix bundle, Residues conserved across the rhoGAP family are largely confined to one face of this bundle, which may be an interaction site for target G protein, In particular, we propose that Arg 85 and Asn 194 are involved in binding G proteins and enhancing GTPase activity.