Alpha4 protection of PP2Ac from degradation requires both the Mid1 binding domain and the PP2Ac binding domain

Protein phosphatase 2A (PP2A) is serine/threonine phosphatase that is regulated through a variety of mechanisms, including post‐translational modifications and association with regulatory proteins. Alpha4 binds to and regulates PP2A catalytic subunit (PP2Ac) stabilizing it through protecting PP2Ac from polyubiquitination and degradation. Alpha4 is a multi‐domain protein with a C‐terminal domain that binds Mid1, an E3 ubiquitin ligase, and an N‐terminal domain containing the PP2Ac binding site and a UIM consensus motif, shown to be important in protecting PP2Ac from polyubiquitination. We will present the structure of the N‐terminal domain of alpha4 determined by x‐ray crystallography and show that it is a flexible tetratricopeptide repeat‐like protein. We also demonstrate that both the Mid1 binding domain and the PP2Ac binding domain of alpha4 are essential to protect PP2Ac from degradation and polyubiquitination. The actual mechanism by which alpha4 protects PP2Ac from degradation appears to involve contributions from multiple domains of alpha4 indicating a multi‐part mechanism of action that has yet to be fully elucidated.