Phosphatase and tensin homolog is a lipid phosphatase that serves as the major negative regulator of the PI3K/AKT pathway. It catalyzes the 3′-specific dephosphorylation of phosphatidylinositol (3,4,5)-trisphosphate (PIP₃) to generate PIP₂. PTEN's lipid phosphatase function depends on several domains, including an N-terminal segment spanning the first 24 amino acids, which results in a catalytically impaired enzyme when mutated. Furthermore, PTEN is regulated by a cluster of phosphorylation sites located on its C-terminal tail at Ser380, Thr382, Thr383, and Ser385, which drives its conformation from an open to a closed autoinhibited but stable state. Herein, we discuss the protein chemical strategies we used to reveal the structure and mechanism of how PTEN's terminal regions govern its function.