Identification of the Mg2+-binding site in the P-type ATPase and phosphatase members of the HAD (haloacid dehalogenase) superfamily by structural similarity to the response regulator protein CheY

Abstract

The large HAD (haloacid dehalogenase) superfamily of hydrolases comprises P-type ATPases, phosphatases, epoxide hydrolases and L-2-haloacid dehalogenases. A comparison of the three-dimensional structure of L-2-haloacid dehalogenase with that of the response regulator protein CheY allowed the assignment of a conserved pair of aspartate residues as the Mg2+-binding site in the P-type ATPase and phosphatase members of the superfamily. From the resulting model of the active site, a conserved serine/threonine residue is suggested to be involved in phosphate binding, and a mechanism comprising a phosphoaspartate intermediate is postulated.

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“…Analogies between interactions in the active sites of BeF 3 Ϫ ⅐PSP and activated CheY (Fig. 3C) provide evidence that Thr-625 and Lys-684, respectively, of the Ca 2ϩ -ATPase form the corresponding hydrogen bond and salt bridge to the phosphoaspartate, as previously postulated (29). In the structure of the unphosphorylated calciumbound form of the ATPase that has been determined (32), Lys-684 appears to form a salt bridge to Asp-351 that is similar to the salt bridge between Lys-109 and Asp-57 in inactive CheY (44).…”

“…High-resolution crystal structures of HAD superfamily members [haloacid dehalogenases from Pseudomonas sp. YL and Xanthobacter autotrophicus GJ10 (30,31), the Ca 2ϩ -ATPase from rabbit sarcoplasmic reticulum (32), and PSP from Methanococcus jannaschii (33)] indicate that the catalytic domains of these proteins form a typical ␣͞␤ Rossmann fold that is similar to that in CheY (29). The quintet of conserved residues noted above surrounds the active site, as it does in receiver domains.…”

“…The quintet of conserved residues noted above surrounds the active site, as it does in receiver domains. These commonalities led to proposals of a common reaction mechanism for the formation of the phosphoaspartate bond (12,29). They also raise the question of whether BeF 3 Ϫ might yield a persistent aspartyl phosphate analog in members of the HAD superfamily and thus facilitate structural studies of their conformations during catalysis.…”

“…The core catalytic domains of HAD family phosphotransferases and P-type ATPases share the same quintet of highly conserved residues found in receiver domains (12,29), although a circular permutation is required to align the primary sequences of members of these two large families (29). High-resolution crystal structures of HAD superfamily members [haloacid dehalogenases from Pseudomonas sp.…”

Insulin Controls Subcellular Localization and Multisite Phosphorylation of the Phosphatidic Acid Phosphatase, Lipin 1

“…Imbedded in the CLIP domain is a signature sequence of the HAD superfamily of hydrolases (12). In HAD family phosphatases, the phosphate group from the substrate is transferred to Asp (corresponding to Asp 712 in lipin 1b) in the HAD motif I consensus sequence, ⌽⌽⌽⌽DXXGT⌽ (where ⌽ represents a bulky hydrophobic residue and X is any residue) (13).…”