The phosphoinositide 3-kinase (PI3K) superfamily includes lipid kinases (PI3Ks and type III PI4Ks) and a group of PI3K-like Ser/Thr protein kinases (PIKKs: mTOR, ATM, ATR, DNA-PKcs, SMG1 and TRRAP) that have a conserved C-terminal kinase domain. A common feature of the superfamily is that they have very low basal activity that can be greatly increased by a range of regulatory factors. Activators reconfigure the active site, causing a subtle realignment of the N-lobe of the kinase domain relative to the C-lobe. This realignment brings the ATP-binding loop in the N-lobe closer to the catalytic residues in the C-lobe. In addition, a conserved C-lobe feature known as the PIKK regulatory domain (PRD) also can change conformation, and PI3K activators can alter an analogous PRD-like region. Recent structures have shown that diverse activating influences can trigger these conformational changes, and a helical region clamping onto the kinase domain transmits regulatory interactions to bring about the active site realignment for more efficient catalysis. A recent report of a small-molecule activator of PI3Kα for application in nerve regeneration suggests that flexibility of these regulatory elements might be exploited to develop specific activators of all PI3K superfamily members. These activators could have roles in wound healing, anti-stroke therapy and treating neurodegeneration. We review common structural features of the PI3K superfamily that may make them amenable to activation.