2,3-Diphosphoglycerate phosphatase activity of phosphoglycerate mutase: stimulation by vanadate and phosphate

Stankiewicz, Paul J.; Gresser, Michael J.; Tracey, Alan S.; Hass, Louis F.

doi:10.1021/bi00379a010

Cited by 49 publications

(31 citation statements)

References 15 publications

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“…6c was obtained Fig. 6 is consistent with a 1:1:1 stoichiometry of a ternary complex of alendronate, metal ion, and H 2 O 2 as the inactivating species (30), although the three components could also be acting synergistically to inactivate the enzyme without forming a ternary complex.…”

Section: Fig 4 Elimination Of Inhibition Of Cd45supporting

confidence: 74%

How Does Alendronate Inhibit Protein-tyrosine Phosphatases?

Skorey¹,

Ly²,

Kelly³

et al. 1997

Journal of Biological Chemistry

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Alendronate(4-amino-1-hydroxybutylidene1,1-bisphosphonate) is a drug used in the treatment of osteoporosis and other bone diseases. The inhibition of proteintyrosine phosphatases (PTPs) by alendronate suggests that PTPs may be molecular targets. As a clear understanding of the inhibition mechanism is lacking, our aim was to analyze the mechanism to provide further insight into its therapeutic effect. We show here that the inhibition of PTPs by alendronate in the presence of calcium followed first-order kinetic behavior, and kinetic parameters for the process were determined. Evidence is presented that the inhibition by alendronate/calcium is active site-directed. However, this process was very sensitive to assay constituents such as EDTA and dithiothreitol. Furthermore, the inhibition of PTPs by alendronate/calcium was eliminated by the addition of catalase. These observations suggest that a combination of alendronate, metal ions, and hydrogen peroxide is responsible for the inhibition of PTPs. The individual effects of alendronate, calcium, or hydrogen peroxide on the inactivation of CD45 were determined. Electrospray ionization mass spectrometry demonstrated that the mass of PTP1B increased by 34 ؎ 2 units after the enzyme was inactivated with alendronate/calcium, due to the oxidization of the catalytic cysteine to sulfinic acid (Cys-SO 2 H). The inhibited PTP1B could be partially reactivated by treatment with reducing agents such as hydroxylamine (NH 2 OH) and N,N-dimethyl-N,N-bis-(mercaptoacetyl)hydrazine, indicating the presence of other oxidized forms such as sulfenic acid (Cys-SOH). This further confirms that the inhibition is the result of oxidation of the catalytic cysteine. The relevance of this oxidative inhibition mechanism in a biological system is discussed.Alendronate (4-amino-1-hydroxybutylidene 1,1-bisphosphonate) has been successfully used as a therapeutic agent for the treatment of bone disorders such as osteoporosis (1-4) and Paget's disease (5, 6). The rationale for the therapeutic effect at the cellular level is that alendronate interferes with osteoclast resorption, resulting in reduced bone turnover and a net gain in bone mineral density (7). However, at the molecular level, the precise target and mechanism of action of alendronate are not yet clear. The evidence that vanadate, which is a potent and nonselective protein-tyrosine phosphatase (PTP) 1 inhibitor (8), inhibits bone resorption induced by parathyroid hormone in organ culture (9) suggests that PTP is possibly one of the targets.Protein-tyrosine phosphatases are important enzymes in living systems, acting in opposition to protein-tyrosine kinases to control the tyrosine phosphorylation level of proteins. Tyrosine phosphorylation plays a critical role in the transduction of signals that control many diverse cell processes (10 -12). PTPs are divided into two main groups: the receptor-like forms, such as CD45 (leukocyte common antigen); and the cytoplasmic forms, such as PTP1B. Each of the PTPs contains at least one conserved segment...

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Section: Fig 4 Elimination Of Inhibition Of Cd45supporting

confidence: 74%

How Does Alendronate Inhibit Protein-tyrosine Phosphatases?

Skorey¹,

Ly²,

Kelly³

et al. 1997

Journal of Biological Chemistry

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“…15,16 The broadening of NMR vanadate signals upon interaction with proteins has also been previously reported for several vanadate complexes. [17][18][19][20] As observed by SDS-PAGE gel electrophoresis, the isolation of actin, according to Pardee and Spudich, 21 produced a G-actin (42.3 kDa) with 99% purity (not shown), which allows concluding that the NMR observations described above are due to the interaction of the monomeric state of actin, G-actin, with decavanadate. On the other hand, the addition of actin in its polymerized form, F-actin, up to 50 mM, induces a much smaller broadening of V 10 signals (1.5-fold), in comparison with G-actin, suggesting that decavanadate protein binding sites are encrusted upon actin polymerization.…”

Section: Introductionmentioning

confidence: 76%

Recent advances into vanadyl, vanadate and decavanadate interactions with actin

2012

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Although the number of papers about ''vanadium'' has doubled in the last decade, the studies about ''vanadium and actin'' are scarce. In the present review, the effects of vanadyl, vanadate and decavanadate on actin structure and function are compared. Decavanadate 51 V NMR signals, at À516 ppm, broadened and decreased in intensity upon actin titration, whereas no effects were observed for vanadate monomers, at À560 ppm. Decavanadate is the only species inducing actin cysteine oxidation and vanadyl formation, both processes being prevented by the natural ligand of the protein, ATP. Vanadyl titration with monomeric actin (G-actin), analysed by EPR spectroscopy, reveals a 1 : 1 binding stoichiometry and a K d of 7.5 mM À1 . Both decavanadate and vanadyl inhibited G-actin polymerization into actin filaments (F-actin), with a IC 50 of 68 and 300 mM, respectively, as analysed by light scattering assays, whereas no effects were detected for vanadate up to 2 mM. However, only vanadyl (up to 200 mM) induces 100% of G-actin intrinsic fluorescence quenching, whereas decavanadate shows an opposite effect, which suggests the presence of vanadyl high affinity actin binding sites. Decavanadate increases (2.6-fold) the actin hydrophobic surface, evaluated using the ANSA probe, whereas vanadyl decreases it (15%). Both vanadium species increased the e-ATP exchange rate (k = 6.5 Â 10 À3 s À1 and 4.47 Â 10 À3 s À1 for decavanadate and vanadyl, respectively). Finally, 1 H NMR spectra of G-actin treated with 0.1 mM decavanadate clearly indicate that major alterations occur in protein structure, which are much less visible in the presence of ATP, confirming the preventive effect of the nucleotide on the decavanadate interaction with the protein. Putting it all together, it is suggested that actin, which is involved in many cellular processes, might be a potential target not only for decavanadate but above all for vanadyl. By affecting actin structure and function, vanadium can regulate many cellular processes of great physiological significance.

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“…The only enzyme known to be activated by VV is phosphoglycerate mutase (17). The two charged vanadate moieties occupy the binding loci for the carboxylate and phosphate of the monophosphoglycerate in the active site of the enzyme.…”

Section: Discussionmentioning

confidence: 99%

“…The chemical formation of phosphovanadyl has been studied previously (16) but not in a biological context. Binding of VV to phosphoglycerate mutase was found to activate the enzyme and the transfer of phosphate directly to a water molecule (17). A crystal structure of VV bound in the active site of the tyrosine phosphatase YopH has been reported recently (18).…”

mentioning

confidence: 99%

Pyrovanadolysis, a Pyrophosphorolysis-like Reaction Mediated by Pyrovanadate, Mn2+, and DNA Polymerase of Bacteriophage T7

Akabayov¹,

Kulczyk²,

Akabayov³

et al. 2011

Journal of Biological Chemistry

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DNA polymerases catalyze the 3-5-pyrophosphorolysis of a DNA primer annealed to a DNA template in the presence of pyrophosphate (PP i ). In this reversal of the polymerization reaction, deoxynucleotides in DNA are converted to deoxynucleoside 5-triphosphates. Based on the charge, size, and geometry of the oxygen connecting the two phosphorus atoms of PP i , a variety of compounds was examined for their ability to carry out a reaction similar to pyrophosphorolysis. We describe a manganese-mediated pyrophosphorolysis-like activity using pyrovanadate (VV) catalyzed by the DNA polymerase of bacteriophage T7. We designate this reaction pyrovanadolysis. X-ray absorption spectroscopy reveals a shorter Mn-V distance of the polymerase-VV complex than the Mn-P distance of the polymerase-PP i complex. This structural arrangement at the active site accounts for the enzymatic activation by Mn-VV. We propose that the Mn 2؉ , larger than Mg 2؉ , fits the polymerase active site to mediate binding of VV into the active site of the polymerase. Our results may be the first documentation that vanadium can substitute for phosphorus in biological processes.Gene 5 of bacteriophage T7 encodes a DNA polymerase essential for replication of the T7 genome (1). T7 DNA polymerase forms a complex with Escherichia coli thioredoxin (trx), 2 an interaction that increases its processivity of nucleotide polymerization several hundredfold (2). The structure of T7 DNA polymerase in complex with trx is shown in Fig. 1. The structure closely resembles that of E. coli DNA polymerase I, thus placing it in the polymerase I family of DNA polymerases (3). In this study, we designate the polymerase in complex with trx as T7 DNA polymerase. Like most other prokaryotic DNA polymerases, T7 DNA polymerase has a proofreading 3Ј-5Ј-exonuclease activity located in the amino-terminal half of the protein (3). The crystal structure shows that its active site is located between the "fingers" and the "palm" subdomains and contains two magnesium ions (Mg 2ϩ ). During polymerization of nucleotides, the Mg 2ϩ closest to the primer (catalytic Mg 2ϩ ) is responsible for deprotonation of the 3Ј-hydroxyl group of the primer prior to its nucleophilic attack on the ␣-phosphate of the incoming dNTP. The second Mg 2ϩ (structural Mg 2ϩ ) is associated with stabilization of the reactive state of the ␤-and ␥-phosphates of the incoming deoxynucleoside 5Ј-triphosphate (dNTP) (4).In the presence of PP i , DNA polymerases catalyze the 3Ј-5Ј-degradation of a DNA primer annealed to a DNA template (5). In this reaction, known as pyrophosphorolysis, the deoxynucleotides (dNMP) of the DNA are converted to dNTP. Pyrophosphorolysis is a true reversal of the polymerization reaction in that the products are dNTPs, and it has the same requirements for a template and a 3Ј-hydroxyl-terminated primer (5). The 3Ј-5Ј-exonuclease associated with DNA polymerase, however, catalyzes the 3Ј-5Ј hydrolysis of the primer to yield dNMPs without a strict requirement for proper base pairing. Pyrophosphorolysis o...

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2,3-Diphosphoglycerate phosphatase activity of phosphoglycerate mutase: stimulation by vanadate and phosphate

Cited by 49 publications

References 15 publications

How Does Alendronate Inhibit Protein-tyrosine Phosphatases?

How Does Alendronate Inhibit Protein-tyrosine Phosphatases?

Recent advances into vanadyl, vanadate and decavanadate interactions with actin

Pyrovanadolysis, a Pyrophosphorolysis-like Reaction Mediated by Pyrovanadate, Mn2+, and DNA Polymerase of Bacteriophage T7

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