Molecular model of the G protein alpha subunit based on the crystal structure of the HRAS protein.

Abstract

A structural model of guanine nucleotide-binding regulatory protein .alpha. subunits (G.alpha. subunits) is proposed based on the crystal structure of the catalytic domain of the human HRAS protein (p21rAs). Because of low overall sequence similarity, structural and functional constraints were used to align the G.alpha. consensus sequence with that of p21ras. The resulting G.alpha. model specific the spatial relationship among the guanine nucleotide-binding site, the binding site of the .beta..gamma. subunit complex, likely regions of effector and receptor interaction, and sites of cholera and pertussis toxin modifications. The locations in the model of the experimentally determined sites of proteolytic digestion, point mutation, and monoclonal antibody binding, and toxin modification are consistent with and help explain the observed biological activity. Two important findings from our model are (i) the orientation of the G.alpha. model with respect to the membrane and (i) the identification of the spatial proximity of the N- and C-terminal regions. Furthermore, the analogy to p21ras, the model assigns specific residues in G.alpha. required for binding the guanosine (G-box) and phosphates (PO4-box) and identifies residues potentially involved in the conformational switch mechanism (S-box). Specification of these critical regions in the G.alpha. model suggests guidelines for construction of mutants and chimeric proteins to experimentally test structural and functional hypotheses.