Crystal structures of rat acid phosphatase complexed with the transition‐state analogs vanadate and molybdate

Abstract

The three-dimensional structures of complexes of recombinant rat prostatic acid phosphatase with the transition-state analogs vanadate and molybdate were determined to 0.3-nm resolution using protein crystallographic methods. The overall structure of the enzyme remains unchanged upon binding of the metal oxyanions; only local conformational differences in the positions of some side chains at the active site were found. The metal oxyanions bind in an identical fashion at the active site with trigonal bipyramidal coordination geometry. The metal ion is within coordination distance of the His12 side chain which is located at one of the axial positions. The three equatorial oxygen atoms interact with the conserved residues Argll, Argl.5, Arg79 and His257. Within hydrogen-bonding distance of the axial oxygen atom is the side chain of the conserved residue Asp258. The implications of these results for the catalytic mechanism of acid phosphatase are discussed.The hydrolysis of phosphoric monoesters is catalyzed by a number of different, non-related enzymes. High-molecularmass acid phosphatases (ortho-phosphoric-monoester phosphohydrolases) form one class of phosphoric-monoester phosphatases which also catalyze phosphoryl transfer between a phosphoester and alcohols (Bodansky, 1972;Van Etten, 1982;Vincent et al., 1992). Alignment of amino acid sequences of acid phosphatases (Roiko et al., 1990;Van Etten et al., 1991) has revealed a conserved sequence motif RHGXRXP. The RHG sequence is also found in phosphoglycerate mutase and 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase which suggested that these three enzymes might share a common fold (Bazan et al., 1989). Recent crystallographic studies Lindqvist et al., 1993) of native rat prostatic acid phosphatase and its complex with the inhibitor L(+)-tartrate have shown that the structure of acid phosphatase consists of two domains, an utP domain consisting of a seven-stranded mixed P-sheet with a-helices on both sides of the sheet, and a smaller ahelical domain. The overall fold of the alp domain is indeed similar to the fold observed in phosphoglycerate mutase. The active site is located at the C-terminus of the P-strands in the alp domain and the conserved residues Argll, Hisl2, Argl5, Arg79 are part of the phosphate-binding site. In the binary complex with the inhibitor L( +)-tartrate, these residues and the conserved residues His257 and Asp258 are involved in binding of the inhibitor. Site-directed mutagenesis studies of acid phosphatase from Escherichia coli showed that the residues corresponding to Argll and His12 are required for catalysis and that substitutions of residues corresponding to Arg15, Arg79, His257 and Asp258 severely impair the catalytic activity (Ostanin et al., 1992;Ostanin and Van Etten, 1993) consistent with the location of these residues at the active site. The chemical pathway of phosphoric-ester hydrolysis by acid phosphatases is summarized in Scheme 1 and is supported by many studies (Van Etten, 1982;Vincent et al., 1992). The hydrolysis can...

Computational Modeling and Experimental Evaluation of a Novel Prodrug for Targeting the Extracellular Space of Prostate Tumors

“…2B). The phosphorus atom is at a favorable distance for nucleophile attack by His 12 , leading to the dephosphorylation of IQ 2-P to IQ 2-OH , in agreement with the theoretical axial attack of P by this nucleophile (24,25). The iodine atom of the quinazolinone points outward from the active site and does not hinder the binding to PAP.…”

“…2A) 18 and Trp 174 as in the case of the crystallized complex. Furthermore, the docking position of the phosphorus atom of the phosphonic group of BABPA is close to the crucial His 12 as seen in the crystal structure complex or in other phosphate-based inhibitors (24,25).…”

Crystal Structure of the PTPL1/FAP-1 Human Tyrosine Phosphatase Mutated in Colorectal Cancer

“…To our knowledge, none of the receptor or nonreceptor PTPs reported to date possess a phosphate at the catalytic site. Several PTPs have been crystallized with the phosphate mimetics vanadate and sulfate that are localized at a similar position to the phosphate group on PTPL1 (27)(28)(29)(30). It is believed that the WPD loop in PTPs moves into a "closed" conformation when substrates bind (31).…”

The oxygen isotope composition of phosphate released from phytic acid by the activity of wheat and <i>Aspergillus niger</i> phytase

“…Most plant and fungal phytases belong to the histidine acid phosphatases, which share the same amino acid sequence motif (RHGXRXP) at their active sites as acid phosphatases and nucleotidases (van Etten et al, 1991;Oh et al 2004;Kostrewa et al, 1997Kostrewa et al, , 1999Lim et al, 2000). The amino acid sequence motif at the active site of phosphatases drives the reaction mechanisms, which can lead to either the incorporation of an oxygen atom derived from a water molecule into the newly formed phosphate (Lindqvist et al, 1994;Knöfel and Sträter, 2001;Ortlund et al, 2004), e.g., acid phosphatases or nucleotidases, or to the incorporation of an oxygen atom derived from a hydroxide ion, e.g., alkaline phosphatases (Kim and Wickoff, 1991;Stec et al, 2000). It has been suggested that these two types of reaction mechanisms are the reason why different phosphomonoesterases cause different isotopic fractionations (von Sperber et al, 2014).…”