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. 2017 Jan 26;292(11):4544–4555. doi: 10.1074/jbc.M116.765743

FIGURE 5.

FIGURE 5.

HPLC chromatographs showing net 5-InsP7 kinase activities and InsP8 phosphatase activities of wild-type and phosphatase mutant versions of human PPIP5Ks. Kinase assays, performed as described under “Experimental Procedures” for 120 min with 1 μm 5-InsP7, contained the following: no enzyme (open triangles) (A), 90 ng of either wild-type PPIP5K1 (open circles) or PPIP5K1R399A (closed circles) (B), or 90 ng of either wild-type PPIP5K2 (open circles) or PPIP5K2R388A (closed circles) (C). Assays were quenched, neutralized, and then analyzed by HPLC (see “Experimental Procedures”). Phosphatase assays, performed as described under “Experimental Procedures” for 30 min with 1 μm InsP8, contained the following: no enzyme (open triangles) (D), 25 ng of either wild-type PPIP5K1 (open circles) or PPIP5K1R399A (closed circles) (E), or 40 ng of either wild-type PPIP5K2 (open circles) or PPIP5K2R388A (closed circles) (F). Data shown are representative of three to four biological replicates.