The conformation of bound GMPPNP suggests a mechanism for gating the active site of the SRP GTPase

Background: The signal recognition particle (SRP) is a phylogenetically con served ribonucleoprotein that mediates cotranslational targeting of secrete d and membrane proteins to the membrane. Targeting is regulated by GTP bind ing and hydrolysis events that require direct interaction between structura lly homologous "NG" GTPase domains of the SRP signal recognition subunit an d its membrane-associated receptor, SR alpha. Structures of both the apo an d GDP bound NG domains of the prokaryotic SRP54 homolog, Ffh, and the proka ryotic receptor homolog, FtsY, have been determined. The structural basis f or the GTP-dependent interaction between the two proteins, however, remains unknown. Results: We report here two structures of the NG GTPase of Ffh from Thermus aquaticus bound to the nonhydrolyzable GTP analog GMPPNP. Both structures reveal an unexpected binding mode in which the beta -phosphate is kinked aw ay from the binding site and magnesium is not bound. Binding of the GTP ana log in the canonical conformation found in other GTPase structures is precl uded by constriction of the phosphate binding P loop. The structural differ ence between the Ffh complex and other GTPases suggests a specific conforma tional change that must accompany movement of the nucleotide from an "inact ive" to an "active" binding mode. Conclusions: Conserved side chains of the GTPase sequence motifs unique to the SRP subfamily may function to gate formation of the active GTP bound co nformation. Exposed hydrophobic residues provide an interaction surface tha t may allow regulation of the GTP binding conformation, and thus activation of the GTPase, during the association of SRP with its receptor.