Purple acid phosphatases hydrolysis

Purple acid phosphatases (PAPs) occur widely in nature and are responsible for hydrolysis of orthophosphate monoethers to alcohols under acidic conditions according to the reaction... 

The iron(II)-iron(III) form of purple acid phosphatase (from porcine uteri) was kinetically studied by Aquino et al. (28). From the hydrolysis of a-naphthyl phosphate (with the maximum rate at pH 4.9) and phosphate binding studies, a mechanism was proposed as shown in Scheme 6. At lower pH (ca. 3), iron(III)-bound water is displaced for bridging phosphate dianion, but little or no hydrolysis occurs. At higher pH, the iron(III)-bound OH substitutes into the phosphorus coordination sphere with displacement of naphthoxide anion (i.e., phosphate hydrolysis). The competing affinity of a phosphomonoester anion and hydroxide to iron(III) in purple acid phosphatase reminds us of a similar competing anion affinity to zinc(II) ion in carbonic anhydrase (12a, 12b). 

The active site similarities listed above belie a remarkable functional diversity, which includes phosphate ester hydrolysis, dioxygen and NO reduction, reversible O2 binding, and O2 activation, the last of which includes enzymes involved in ribonucleotide reduction, hydrocarbon monooxygenation, and fatty acyl desaturation. At the overall protein level, the purple acid phosphatases (PAPs) seem to be completely unrelated, both structurally and functionally, to any of the others in this class. Similarly, the flavo-diiron enzymes form a structurally and probably functionally distinct family of proteins, catalyzing both dioxygen and NO reduction. These last two examples illustrate that attempts to shoehorn all of these enzymes into a single class can sometimes provide a simplistic and misleading view of their chemistry and biochemistry. 

Purple acid phosphatases (PAPs) catalyze the hydrolysis of phosphate monoesters with mildly acidic pH optima (5-7) utilizing a binuclear metal center containing a ferric ion and a divalent metal ion. PAPs are also characterized by their purple color, the result of a tyrosine (Tyr) to Fe3+ charge transfer transition at about 560nm.113 All known mammalian PAPs are monomeric and have a binuclear Fe3+-Fe2+ center, whereas the kidney bean and soybean enzymes are dimeric and have an Fe3 + -Zn2+ center in each subunit. The X-ray structures for kidney bean PAP114 and the PAP115 from rat bone reveal that despite a sequence similarity of only 18%, they share very similar catalytic sites. The structure of the kidney bean PAP shows the two metal ions at a distance of 3.1 A, with a monodentate bridging Asp-164. These and other residues involved in metal coordination can be seen in Fig. 21. 

The purple acid phosphatases (PAP) catalyze the hydrolysis of phosphate esters under acidic pH conditions (pH optimum 5) (9, 10). They differ from other acid phosphatases in having a distinct purple color due to the presence of iron or manganese and in being uninhibited by tartrate. Diiron units have been found in the active sites of the enzymes from mammalian spleen (171-173) and uterus (173, 174), while a heterodinu-clear FeZn unit has been characterized for the enzyme from red kidney bean (175). Either the Fe2 or the FeZn unit is catalytically competent in these enzymes, since the enzymes from porcine uterus and bovine spleen can be converted into active FeZn forms and the kidney bean enzyme can be transformed into an active Fe2 form (176). There are also enzymes from other plant sources (particularly sweet potato) that have been reported to have either a mononuclear Mn(III) or Fe(III) active site (177), but these are beyond the scope of the review. This section will focus on the enzymes from porcine uterus (also called uteroferrin), bovine spleen, and red kidney bean. 

Purple acid phosphatase Phosphate ester hydrolysis 182)... 

Purple Acid Phosphatases. Purple acid phosphatases (PAPs) utilize a dinuclear metal center to catalyze the hydrolysis of phosphate monoesters. The characteristic purple color of these enzymes arises from a charge transfer absorption at about 560 nm, between a tyrosinate ligand and the conserved Fe + found in all PAPs. The second metal ion varies with the source of the enzyme and is always divalent. Mammalian PAPs are monomeric and have Fe -Fe " centers, whereas most plant PAPs are dimeric with Fe " -Zn + centers. A PAP isolated from sweet potato contains an Fe +-Mn + center, the first of its kind in any enzyme (26,27). This novel PAP also differs from others by its greater catalytic efficiency toward both activated and unactivated substrates (27), as well as in its strict requirement for manganese in the divalent site (26). 

Figure 37. Proposed mechanism of hydrolysis by purple acid phosphatases. Reprinted with permission from [527]. Copyright 2006, American Chemical Society.

Merkx M, Averill BA. 1999. Ifrobing the role of the trivalent metal in phosphate ester hydrolysis preparation and characterization of purple acid phosphatases containing Al Zn and rn Zn active sites, including the first example of an active aluminum enzyme. JAm Chem Soc 121 6683-6689. 

Proteins with dinuclear iron centres comprise some prominent and well studied representatives like ribonucleotide reductase (RNR), purple acid phosphatase (PAP), methane monooxygenase hydroxylase (MMOH), ruberythrin and hemerythrin. The last of these is an oxygen carrier in some sea worms it has been well characterized within this group and has thus laid the foundation to this class of iron coordination motif. Ruberythrin is found in anaerobic sulfate-reducing bacteria. Its name implies that, in addition to a hemerythrin-related diiron site another iron is coordinated in a mononuclear fashion relating to rubredoxin, which is an iron-sulfur centre. The latter will not be treated here. The hydroxylase component of methane monooxygenase is one of the three components in soluble methane monooxygenase (MMO) and contains the active diiron site it is found in methanotropic bacteria. Purple add phosphatase (PAP) occurs mainly in plants and animals, and catalyses the hydrolysis of monophosphate esters. Finally, ribonucleotide reductase reduces ribonucleotides to deoxyribonucleotides and thus has a key position in DNA synthesis. 

Two possible difficulties with the straightforward view that the purple enzymes function as true phosphatases might be considered. First, they display the interesting and poorly understood phenomenon of inhibition by inorganic phosphate, a reaction product of phosphatase activity. The apparent Ki for phosphate acting as a competitive inhibitor of the hydrolysis of p-nitrophenyl phosphate by uteroferrin is 3.2 mM at pH 4.9 Very likely, a similar Kj is exhibited by the tissue phosphatases, in view of their similarities in spectroscopic and structural properties to uteroferrin. Intracellular inorganic phosphate, which may exceed 1 mM in concentration may then act to restrain the phosphatase activity of spleen purple acid phosphatase and its intracellular relatives. 

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