GTPASE MECHANISM OF GPROTEINS FROM THE 1.7-ANGSTROM CRYSTAL-STRUCTURE OF TRANSDUCIN ALPHA-CENTER-DOT-GDP-CENTER-DOT-ALF4(-)

ALUMINIUM fluoride (AlF4-) activates members of the heterotrimeric G-protein (G(alpha beta gamma)) family(1,2) by binding to inactive G(alpha).GDP near the site occupied by the gamma-phosphate in G(alpha).GTP (ref. 3). Here we describe the crystal structure of transducin alpha.GDP activated with aluminium fluoride (G(t alpha).GDP.AlF4-.H2O) at 1.7 Angstrom, a resolution sufficient to establish the coordination geometry of the bound aluminium fluoride as well as the extensive network of direct and water-mediated interactions that stabilize it. These observations are derived from three independent representations in the asymmetric unit, eliminating any chance of drawing conclusions based on stereochemistry imposed by crystal packing. Surprisingly, aluminium fluoride activates G(alpha).GDP by binding with a geometry resembling a pentavalent intermediate for GTP hydrolysis. The stabilizing interactions involve not only residues that interact with the gamma-phosphate in G(t alpha).GTP gamma S, but also conserved residues essential for GTPase activity. Thus the G(t alpha).GDP.AlF4-.H2O structure provides new insight into the mechanism of GTP hydrolysis.