CLONING AND CHARACTERIZATION OF A RHOGAP HOMOLOG FROM DICTYOSTELIUM-DISCOIDEUM

Small GTPases interact with a variety of proteins that affect nucleoti de binding and cleavage. GTPase activating proteins (GAPs) art: one cl ass of these proteins that act by accelerating ire intrinsic GTPase ra re resulting in the formation of the biologically inactive GDP-bound f arm of the GTPase, For the Rho subfamily of GTPases, there is a growin g number of proteins with rhoGAP activity that are identifiable by a h omologous region of about 150 amino acids, We have exploited this homo logy using the polymerase chain reaction to clone the first rhoGAP hom olog, called DdRacGAP, from the slime mold Dictyostelium discoideum. T he GAP domain of DdRacGAP (amino acids 1-212), when expressed and puri fied from Escherichia coli, is active on both Dictyostelium and human Rho Family GTPases but not human Ras. The full-length protein is 1356 amino acids in length and has several interesting homologies ill addit ion to the GAP domain, including an SH3 domain, a dbl homology domain, and a pleckstrin homology domain.