Arf GTPases are the major regulators of most aspects of lipid and membrane traffic. In the resting cell they are GDP-bound. Upon an agonist stimulation of the cell, GEFs promote the exchange of GDP to GTP resulting in an active membrane-bound conformation that interacts specifically with downstream effectors.

To invade their host and avoid being destroyed, intracellular bacterial pathogens inject their effector proteins into the host cytosol. These effectors function as illicit regulators of small GTPases. Legionella pneumophilia, the bacterium that causes severe pneumonia (Legionnaire’s disease), injects a large number of effectors that divert membrane traffic to establish a vacuole where it hides and replicates. One of these effector proteins is RalF, which diverts the function of Arf1.

Ralf molecules are being auto-inhibited in cytosolic environment and need to undergo a major conformational change to reach their active, membrane-bound conformation. Using the recently developed method of Molecular Dynamics with Excited Normal Modes (MDeNM) the mechanism of auto-inhibition release and the membrane-bound open conformation of Ralf are presented in this study.

The mechanism is understood by the conformational mapping with the help of the MDeNM method and its result filtering, based on experience based knowledge. All atom mechanism is presented in this study where the comparison of active and inactive RalF was possible.

Interaction pattern between the GEF-like RalF and small GTPase Arf1 is also described with the identification of crucial residues in both parties. Pivotal residues in RalF’s activation mechanism are revealed.