Multiple conformational switches in a GTPase complex control co-translational protein targeting.

The "GTPase switch" paradigm, in which a GTPase switches between an active, GTP-bound state and an inactive, GDP-bound state through the recruitment of nucleotide exchange factors (GEFs) or GTPase activating proteins (GAPs), has been used to interpret the regulatory mechanism of many GTPases. A notable exception to this paradigm is provided by two GTPases in the signal recognition particle (SRP) and the SRP receptor (SR) that control the co-translational targeting of proteins to cellular membranes. Instead of the classical "GTPase switch," both the SRP and SR undergo a series of discrete conformational rearrangements during their interaction with one another, culminating in their reciprocal GTPase activation. Here, we show that this series of rearrangements during SRP-SR binding and activation provide important control points to drive and regulate protein targeting. Using real-time fluorescence, we showed that the cargo for SRP--ribosomes translating nascent polypeptides with signal sequences--accelerates SRP.SR complex assembly over 100-fold, thereby driving rapid delivery of cargo to the membrane. A series of subsequent rearrangements in the SRP x SR GTPase complex provide important driving forces to unload the cargo during late stages of protein targeting. Further, the cargo delays GTPase activation in the SRP.SR complex by 8-12 fold, creating an important time window that could further improve the efficiency and fidelity of protein targeting. Thus, the SRP and SR GTPases, without recruiting external regulatory factors, constitute a self-sufficient system that provides exquisite spatial and temporal control of a complex cellular process.