The functional and structural roles of residues Gln114 and Glu117 in elongation factor Tu

The effects of substituting residues Gln114 by Glu and Glu117 by Gln, both situated in the vicinity of the guanine-nucleotide-binding pocket, were investigated in the isolated N-terminal domain (G domain) of elongation factor Tu with respect to the binding of the substrate GDP/GTP, GTPase activity and stability. The major change in the interaction with the guanine nucleotides is a lower affinity for GTP and a reduced GTPase activity when Gln114 is substituted by Glu. This mutation also abolishes most of the selective effects on the GTPase activity induced by the different monovalent cations. Substitution of Glu117 by Gln does not affect the interaction with the guanine nucleotides or the GTPase activity of the G domain in an essential way, but it reduces the stability towards denaturation of the G-domain.GDP complex. Our results therefore suggest, that Gln114 is involved in keeping a functional conformation of the guanine-nucleotide-binding pocket, whereas Glu117 participates in the regulation of the overall conformation of the G domain. Neither of these two residues appears to play a role in the actual GTPase mechanism.