Despite belonging to the phosphoserine- and phosphothreonine-specific phosphoprotein phosphatase (PPP) family, Arabidopsis thaliana Rhizobiales-like phosphatase 2 (RLPH2) strongly prefers substrates bearing phosphorylated tyrosine residues. We solved the structures of RLPH2 crystallized in the presence or absence of sodium tungstate. These structures revealed the presence of a central domain that forms a binding site for two divalent metal ions that closely resembles that of other PPP-family enzymes. Unique structural elements from two flanking domains suggest a mechanism for the selective dephosphorylation of phosphotyrosine residues. Cocrystallization with the phosphate mimetic tungstate also suggests how positively charged residues that are highly conserved in the RLPH2 class form an additional pocket that is specific for a phosphothreonine residue located near the phosphotyrosine residue that is bound to the active site. Site-directed mutagenesis confirmed that this auxiliary recognition element facilitates the recruitment of dual-phosphorylated substrates containing a pTxpY motif.

尽管属于磷酸丝氨酸和磷酸苏氨酸特异性磷酸蛋白磷酸酶（PPP）家族，拟南芥根瘤菌样磷酸酶2（RLPH2）强烈优选带有磷酸化酪氨酸残基的底物。我们解决了在钨酸钠存在或不存在下结晶的RLPH2的结构。这些结构揭示了一个中央结构域的存在，该结构域形成了两个二价金属离子的结合位点，该结合位点与其他PPP系列酶的结合位点非常相似。来自两个侧翼结构域的独特结构元件提示了磷酸酪氨酸残基的选择性去磷酸化的机制。与磷酸盐模拟钨酸盐的共结晶还表明，在RLPH2类中高度保守的带正电荷的残基如何形成一个额外的口袋，该口袋对位于结合到活性位点的磷酸酪氨酸残基附近的磷酸苏氨酸残基具有特异性。