Molecular mechanism of the synergistic phosphorylation of phosphatase inhibitor-2. Cloning, expression, and site-directed mutagenesis of inhibitor-2.

Novel phosphorylation‐dependent regulation in an unstructured protein

“…Human I2 T74A and T74E mutant proteins inhibit PP1 in vitro 80 contrary to the expectation from the MD results here. In vitro I2 inhibition of PP1 requires binding of the two proteins and then I2 blocking substrate binding to PP1.…”

A Versatile Mass Spectrometry-Based Method to Both Identify Kinase Client-Relationships and Characterize Signaling Network Topology

“…It has been previously established that mammalian PPI-2 is phosphorylated by multiple PK, and that phosphorylation is important for both selectivity and activity as an inhibitor. , We observed that the situation is similar in the plant-derived system but with additional elaborations. The Thr-residue within the PxT 72 P motif is phosphorylated in mammalian cells by glycogen shaggy-like protein kinase-3 (GSK3) and cyclin-dependent protein kinases 2 (CDC2). − The A. thaliana homologues of these two PK were included in our study (Table S2); however, they did not phosphorylate the “Thr 72 ”-containing peptide (Table S4). In contrast, we observed that AtPPI-2 was phosphorylated at multiple sites by the CK1-like 10, AME3, and Ser PK-like PK (Figure ).…”

“…Currently the precise role of phosphorylation of different amino acids at AtPPI-2 in inhibition of TOPP remains largely unverified; however 3-D modeling and crystallography studies of mammalian PPI-2 provide support for the proposal that phosphorylation of amino acids in the TOPP binding motif may play a role in a conformational change that affects the binding and/or inhibition activity (Figure ). Moreover it has been suggested that the action of one kinase enhances phosphorylation of other residues by different kinases and is required for potentiation of different kinase action . Although the RVxF, PxTPY, and RKxHY motifs are conserved among Arabidopsis and mammalian PPI-2, some features of the mammalian PPI-2 such as the SGILK and SQ motifs at the N-terminal region are believed to be responsible for the TOPP interaction, but these motifs are absent in AtPPI-2 (Figure S6).…”

Regulation of protein phosphatase-1

“…The physiological relevance of this has been questioned, however [79]. I-2 is also phosphorylated at the Ser82, Ser120 and Ser121 residues by CK-2 in vitro [80,81] and in vivo [82,83], but the CK-2 phosphorylations are reported not to affect the phosphatase activity of the complex [80,84]. Possibly phosphorylation may influence the subcellular distribution of I-2 [83].…”