An Unprecedented Fe^III(μ-OH)Zn^II Complex that Mimics the Structural and Functional Properties of Purple Acid Phosphatases

Abstract

Purple acid phosphatases (PAPs) belong to the family of binuclear metallohydrolases and catalyze the hydrolysis of a variety of phosphoester substrates within the pH range of 4-7. 1 They are the only binuclear metallohydrolases where the necessity for a heterovalent active site (Fe III -M II , where M ) Fe, Zn, or Mn) for catalysis has been clearly established. To date, the crystal structures of PAPs from red kidney bean (rkbPAP), 2a rat, 2b,c pig, 2d human, 2e and sweet potato 2f have been reported. In the structure of rkbPAP, 2a the Fe III ion is coordinated by a tyrosine, a histidine, and an aspartate, and a Zn II ion is coordinated by two histidines and an asparagine. The Fe III Zn II ions are bridged by two oxygen atoms, one from the carboxylate group of an aspartate and the other from a modeled µ-(hydr)oxo group. Two oxygen atoms from a µ-1,3 phosphate group complete the coordination spheres of the Zn II and Fe III ions.Despite the availability of detailed structural data, the catalytic mechanism of PAPs remains a matter of controversy. For rkbPAP, a mechanism in which, in the first step of the catalytic cycle, the substrate binds in a monodentate fashion to the Zn II ion has been proposed. 2a The enzyme-substrate complex is oriented in such a way that a terminal Fe III -bound hydroxide can efficiently attack the phosphorus atom of the substrate, leading to the release of the alcohol product. 2a The monodentate binding of the substrate to Zn II is corroborated by the fact that the addition of phosphate to the Fe III Zn II derivative of bovine spleen PAP does not affect the spectroscopic properties of the Fe III ion at the pH of optimal activity (pH 6.5). 3 However, for pig 4a and sweet potato PAP, 2f,4b an alternative mechanism in which the substrate forms a µ-1,3 phosphate complex, thus placing the µ-(hydr)oxo bridge in an ideal position to act as the reaction-initiating nucleophile, has also been proposed.Homo-and heterodinuclear Fe III M II complexes which are capable of reproducing the structural, spectroscopic, and functional properties of PAPs can be very informative to evaluate the mechanism(s) of these metalloenzymes. Recently, we reported on the syntheses, characterization, and phosphatase-like activity of the heterodinuclear [LFe III (µ-OAc) 2 Zn II ] + complex (H 2 L ) 2-bis[{(2-pyridylmethyl)-aminomethyl}-6-{(2-hydroxybenzyl)-(2-pyridylmethyl)}aminomethyl]-4-methylphenol), and we have proposed that upon dissolving the complex in an aqueous solution the dissociation of the carboxylate groups leads to the formation of the catalytically active [(OH)Fe III -(µ-OH)Zn II (OH 2 )] species, 5 similar to that proposed to be present in the active site of rkbPAP. 2a Herein we report the X-ray structure, solution studies, and phosphatase activity of the first mixed-valence complex containing the Fe III (µ-OH)Zn II motif (1). The molecular structure of 1 (Figure 1) shows that in the dinuclear [L(OH 2 )Fe-(µ-OH)Zn] 2+ unit the Fe III ion is facially coordinated by the hard tridentate pendant arm of L 2-...

Spectroscopic and mechanistic studies of dinuclear metallohydrolases and their biomimetic complexes

“…26 The phosphatase activity of this complex yielded a bell-shaped, strongly pH-dependent rate profile with a pH-optimum at 6.5, and pK a values of 5.3 and 8.1, demonstrating that this complex is an excellent structural and functional model of the active site of PAPs. 26 In order to generate structurally more relevant biomimetics for dinuclear metallohydrolases much effort has been devoted to the synthesis of asymmetric ligands considered to be more suitable models for the asymmetric coordination environment found in enzymatic systems. Nordlander et al proposed that asymmetric complexes are not only more appropriate functional models for the active site of phosphoesterase enzymes, but also that they exhibit enhanced catalytic rates compared with their symmetric counterparts.…”

New La(III) Complex Immobilized on 3-Aminopropyl-Functionalized Silica as an Efficient and Reusable Catalyst for Hydrolysis of Phosphate Ester Bonds

“…Data from the literature, in which dinuclear Ni II Ni II and Fe III Zn II biomimetics and the enzyme Fe III Zn II purple acid phosphatase have been used as the catalysts in the hydrolysis of mono-and diester-phosphate bonds are in full agreement with this proposal. 8, 25,26 For 1 and APS-1, 9 and 13 turnovers in 24 h were detected, providing evidence that during the catalytic cycle (excess of diester), the intermediate containing the monoester dissociates to regenerate the catalytically active species. Indeed, such a conclusion is reinforced by the fact that addition of 1 equiv of the monoester 2,4-DNPP (as a possible inhibitor) to the reaction mixture of APS-1 and 2,4-BDNPP does not affect the number of turnovers in 24 h. Therefore, under excess substrate (2,4-BDNPP), hydrolysis of the monoester to form inorganic phosphate and 2,4-dinitrophenolate occurs mainly due to the background reaction (spontaneous hydrolysis).…”

Phosphate ester cleavage promoted by a tetrameric iron(III) complex

“…Although it is believed that PAPs are only active in the heterovalent state [16], phosphatase activity has been reported for a number of diferric model complexes [17][18][19][20][21][22][23] [8,9,[17][18][19][20][21][22][23][24][25], particularly those for which more than two deprotonation equilibria are considered as part of the profile [9,13,14].…”