(A) Model of SHIP2 Ptase-C2 docked to the membrane and with PI(3,4,5)P3-Mg2+ bound to the active site. Basic and hydrophobic side chains of K531, I534, L538, K568, R571, R581, R588 and L590 in the Ptase and R762, K764, K779, K826 and R859 in the C2 domain (shown as sticks) are modelled to contribute to membrane interactions. L2, containing I534 and L538 is modeled to penetrate the lipid bilayer. The C2-PS interaction is based on PDB entry 1DSY and PI(3,4,5)P3 interactions on the model shown in Figure 6A. (B) The catalytic cycle. Top: Docking of Ptase-C2 to the membrane orients via the rigid domain interface the Ptase active site towards its membrane substrate. Step 1: L4 is ‘out’ to allow entry of the PI(3,4,5)P3 headgroup. Step 2: Initial engagement of the substrate 4 P with N684 destabilizes the doubly bound L4-out conformation and initiates the switch to L4-in, which allows R682 interactions with the substrate 3 P. L2 opens and penetrates the membrane to interact with PI(3,4,5)P3 lipid chains. Step 3: With the substrate correctly positioned, catalysis proceeds and the cleaved 5 P and the Mg2+ion move towards the metal B-site (see text). Step 4: Product interactions are weakened by L4 moving ‘out’, releasing R682 from the product 3 P. Further, N684 releases 4 P interactions by switching to bind H674 (see Figure 1F). Step 5: The weakly bound product is released and a new substrate can be engaged.