Extended Data Fig. 7: PIP2 binding is required for LPP1 function.
From: Structural basis for the catalytic mechanism of human lipid phosphate phosphatases
a, EM maps of PIP2 molecules in these four structures. The densities are contoured at 5σ in the LPP1-R, LPP1-C2M, and LPP1-C3M structures, and at 3σ in the LPP1-C3M structure. The PIP2 molecules are represented as black sticks and balls. b, SEC profiles of enzymes with mutations on essential residues involved in PIP2 coordination. c, Functional verification of essential residues involved in PIP2 coordination using PA and S1P as substrates. Data are presented as the means ± s.d. of three independent experiments, each with two technical repeats (n=6). Significant differences were determined with one-way ANOVA with Tukey’s multiple comparisons tests. d, SDS-PAGE analysis of LPP1 variants in panel c and Fig. 6e. CB, Coomassie blue staining; WB, immunoblotting analysis with Strep antibody. The representative blots shown here are from more than three independent experiments. e, Mechanistic model of LPP1-mediated dephosphorylation of LPA. LPP1 catalyzes the dephosphorylation of LPA via a ‘ping-pong’ mechanism. Step 1: the C2 histidine (His171) facilitates the cleavage of the phosphate bond, and the C3 histidine acts as a nucleophile to form a phosphohistidine intermediate with the cleaved phosphate. Subsequently, the first product, MAG, is released. Step 2: a water molecule enters the catalytic cavity and hydrolyzes the phosphohistidine intermediate, resulting in the release of the 2nd product, inorganic phosphate. The structures we present here likely capture the major intermediate states of Step 1, from substrate binding to the formation of the phosphohistidine intermediate.
