Characterization of a plant, tyrosine-specific phosphatase of the aspartyl class.

The Arabidopsis thaliana homologue of the Eyes Absent genes (AtEYA) encodes a protein corresponding to the C-terminal conserved domain of the animal Eyes Absent proteins. We show here that AtEYA is a tyrosine-specific phosphatase that hydrolyzes its substrates in a metal-dependent reaction analogous to the phosphoserine phosphatases of the haloacid dehalogenase (HAD) family. The animal Eyes Absent proteins are a novel family of dual-function enzymes: they are transcription factors as well as phosphatases. They also represent a new mechanistic class of tyrosine phosphatases (PTPs) that do not have the Cys-containing signature motif. In contrast, AtEYA is only a tyrosine phosphatase and has no transactivation domain. Using the reaction mechanism of other HAD family enzymes as a model, we have conducted mutational analyses on AtEYA to query the roles of conserved residues. This analysis confirms the importance of the putative nucleophile, the general acid, and the metal-binding residues. Additionally, an inhibitory profile that is diagnostic of this new class of protein tyrosine phosphatases is described. The results of these studies on AtEYA reveal that while the animal and plant Eyes Absent proteins catalyze the same dephosphorylation reaction, the details of their specificity and active site environment, as well as their biological roles, are distinct.