Allosteric regulation of the Golgi-localized PPM1H phosphatase by Rab GTPases modulates LRRK2 substrate dephosphorylation in Parkinson's disease.

PPM1H phosphatase reverses Parkinson's disease-associated, leucine-rich repeat kinase 2 (LRRK2)-mediated, Rab GTPase phosphorylation. We showed previously that PPM1H relies on an N-terminal amphipathic helix for Golgi membrane localization, and this helix enables PPM1H to associate with liposomes in vitro; binding to highly curved liposomes activates the phosphatase activity of PPM1H. We show here that PPM1H also contains an allosteric binding site for its nonphosphorylated reaction products, Rab8A and Rab10. Microscale thermophoresis revealed that PPM1H binds thiophosphorylated Rab8A at the active site with a K(D) of ∼1 μM; binding of Rab8A and Rab10 to an alternative site is of similar affinity and is not detected for another LRRK2 substrate, Rab12. Nonphosphorylated Rab8A or Rab10 inhibit PPM1H phosphatase reactions at concentrations consistent with their measured binding affinities and fail to inhibit PPM1H L66R phosphatase reactions. Independent confirmation of nonphosphorylated Rab binding to PPM1H was obtained by sucrose gradient coflotation of nonphosphorylated Rabs with liposome-bound PPM1H. Finally, Rab8A or Rab10 binding also requires PPM1H's amphipathic helix, without which the interaction affinity is decreased about sixfold. These experiments indicate that Golgi-associated Rab proteins contribute to the localization of PPM1H, and nonphosphorylated Rabs regulate PPM1H phosphatase activity via an allosteric site. Targeting this site could represent a strategy to enhance PPM1H-mediated dephosphorylation of LRRK2 substrates, offering a potential therapeutic approach to counteract LRRK2-driven Parkinson's disease.