mTORC2 phosphorylates the protein kinase Akt at Ser473 in the hydrophobic motif after recruitment by PIP3, which induces a conformational change. In cells, mTORC2 has high specificity: it phosphorylates Akt and PKC, but not related kinases, which are substrates of mTORC1. The researchers created semi-synthetic probes to capture the mTORC2-Akt complex and determined its structure using cryo-EM. Akt's local sequence contributes little to mSin1 substrate recognition, instead specificity is determined by secondary and tertiary structural elements of Akt binding to mSin1 distal to the mTOR active site. These elements are conserved in at least 18 related substrates. mTORC2 directly and specifically phosphorylates Ser473 of Akt with minimal local sequence contribution; mutation of Thr443 reduces phosphorylation by only 50%. The Akt PH domain is not essential for C-terminal phosphorylation of Akt mTORC2 in solution, where kcat/KM is less than 2-fold different from wild type. The structure reveals a "substrate arc" where the kinase component Akt interacts with mTORC2, including previously unseen parts of the mSin1 CRIM domain and the mTOR helix.[1][2]