Phosphate Analogues Research Articles

Tf2O-Promoted Activating Strategy of Phosphate Analogues: Synthesis of Mixed Phosphates and Phosphinate.

A metal-, toxic chloride reagent-free activating strategy of various phosphates has been developed. This method enables the facile synthesis of functional phosphates such as alkyl phosphates, aza phosphates, thiophosphate, and mixed diaryl phosphates. A transient phosphorylpyridin-1-ium species in situ generated from phosphates with Tf2O/pyridine readily undergoes a substitution reaction with diverse nucleophiles to form versatile phosphate compounds.

Sodium orthovanadate overcomes sorafenib resistance of hepatocellular carcinoma cells by inhibiting Na+/K+-ATPase activity and hypoxia-inducible pathways

The resistance to sorafenib highly affects its clinical benefits for treating hepatocellular carcinoma (HCC). Sodium orthovanadate (SOV) is a phosphate analog that displays anti-cancer activities against various types of malignancies including HCC. The present study has demonstrated that SOV is able to overcome sorafenib resistance and strengthens sorafenib in suppressing sorafenib-resistant HCC cells in vitro and in animal models. Similar to its action on parental HCC cells, SOV induced cell cycle arrest at G2/M phases by regulating cyclin B1 and cyclin-dependent kinase 1, and apoptosis by reducing mitochondrial membrane potential, in sorafenib-resistant HCC cells. More importantly, SOV inhibited ATPase activity, which was significantly elevated in sorafenib-resistant HCC cells. SOV also reduced the expression of HIF-1α and HIF-2α and their nuclear translocation, resulting in downregulation of their downstream factors including vascular endothelial growth factor, lactate dehydrogenase-A and glucose transporter 1. Its ability to inhibit ATPase activity and hypoxia-inducible pathways enabled SOV to efficiently suppress both normoxic and hypoxic cells, which compose cancer cell populations inside sorafenib-resistant HCC tumors. The present results indicate that SOV may be a potent candidate drug for overcoming the resistance to sorafenib in treating HCC.

Investigation of lead bioimmobilization and transformation by Penicillium oxalicum SL2

Fungi Penicillium oxalicum SL2 was applied for Pb2+ bioremediation in aqueous solution in this study. After 7 days of incubation at different initial concentrations of Pb2+ (0, 100, 500 and 2500 mg L−1), most of Pb2+ were removed (90, 98.3, and 86.2%), the maximum Pb content in mycelium reached about 155.6 mg g−1 dw. Meanwhile, the formation of extracellular secondary minerals and intracellular Pb-complex were observed and identified, the speciation of Pb in mycelium was also detected by X-ray absorption near-edge structure (XANES) spectroscopy, i.e., Pb-oxalate, Pb-citrate, Pb-hydrogen phosphate and Pb-glutathione analogues. In addition, content of glutathione and oxidized glutathione was increased under the exposure of Pb2+, which implied that glutathione might play a key role in Pb immobilization and detoxification in P. oxalicum SL2. This study elucidated partial mechanisms of Pb immobilization and speciation transformation of this strain, providing an alternative biomaterial in the bioremediation of Pb-contaminated wastewater.

Phosphate ester cleavage by a positively charged porous silica adorned with lanthanum (III)

Phosphate esters and analogs thereof are well known in chemistry and biology by their formidable stability. We used a porous silica material (PSiM) functionalized with amino groups (8.4%) and lanthanum ions (1.5%) to catalyze the cleavage of bis(2,4-dinitrophenyl)phosphate (BDNPP). The functionalized PSiM attracted the substrate from solution and accelerated its hydrolysis by 38-fold in relation to the uncatalyzed reaction at pH 8.14. After an induction period associated to hydration of the catalyst during the first use, the catalytic activity increased in the first reuse to about 600-fold. Supramolecular effects were proposed to drive the catalytic process. Kinetic studies in different pH's showed that the lanthanum ion and amino groups acted mutually in the BDNPP hydrolysis and a reaction mechanism that took this into account was proposed.

Phosphate starvation response controls genes required to synthesize the phosphate analog arsenate.

Environmental arsenic poisoning affects roughly 200 million people worldwide. The toxicity and mobility of arsenic in the environment is significantly influenced by microbial redox reactions, with arsenite (AsIII ) being more toxic than arsenate (AsV ). Microbial oxidation of AsIII to AsV is known to be regulated by the AioXSR signal transduction system and viewed to function for detoxification or energy generation. Here, we show that AsIII oxidation is ultimately regulated by the phosphate starvation response (PSR), requiring the sensor kinase PhoR for expression of the AsIII oxidase structural genes aioBA. The PhoRB and AioSR signal transduction systems are capable of transphosphorylation cross-talk, closely integrating AsIII oxidation with the PSR. Further, under PSR conditions, AsV significantly extends bacterial growth and accumulates in the lipid fraction to the apparent exclusion of phosphorus. This could spare phosphorus for nucleic acid synthesis or triphosphate metabolism wherein unstable arsenic esters are not tolerated, thereby enhancing cell survival potential. We conclude that AsIII oxidation is logically part of the bacterial PSR, enabling the synthesis of the phosphate analog AsV to replace phosphorus in specific biomolecules or to synthesize other molecules capable of a similar function, although not for total replacement of cellular phosphate.

Structure-Dependent in Vitro Metabolism of Alkyl-Substituted Analogues of Triphenyl Phosphate in East Greenland Polar Bears and Ringed Seals

Organophosphate esters (OPEs), used as plasticizers and flame retardants, are major emerging environmental contaminants in the Arctic. OPEs of environmental interest include triphenyl phosphate (TPHP) and a growing array of alkyl-substituted TPHP analogues. Using a microsomal assay of the liver tissue of polar bears and their ringed seal prey, the comparative in vitro metabolism of TPHP was investigated relative to the analogues of isodecyl diphenyl phosphate (IDDPP), (p-tert-butylphenyl) diphenyl phosphate (TBPDPP), tris(p-tert-butylphenyl) phosphate (TTBPP), and two tris(isopropylphenyl) phosphate isomers (T2IPPP and T4IPPP). Polar bear metabolism of the p-tert-butylphenyl-substituted OPEs, TBPDPP and TTBPP, had a substantially slower rate and percent metabolic depletion compared to those with TPHP. Isodecyl- and isopropyl-substituted OPEs, IDDPP, T2IPPP, and T4IPPP, were also more slowly depleted by polar bears than TPHP was. TPHP, IDDPP, T2IPPP, TBPDPP, TTBPP, and T4IPPP were all slowly metabolized by...

Computational Study of Butyrylcholinesterase Inhibition by Aryl Alkyl Cholinyl Phosphorus Derivatives

Acetylcholine (ACh) is a neurotransmitter that is released from nerve cells to send signals to other cells, and acetylcholinesterase (AChE) is a highly specific enzyme that hydrolyzes ACh to regulate intercellular communication. However, butyrylcholinesterase (BChE), a non-specific AChE imposter, also breaks down choline based esters such as ACh. The activity of BChE has been found to gradually increase over time in patients with Alzheimer's disease (AD), which has been linked to a significant decrease in synaptic ACh levels, thus disrupting intercellular communication. In the treatment of AD, discovering inhibitors that prevent the activity of BChE and that do not interfere with the activity of AChE is vital. Aryl alkyl cholinyl phosphorous (AACP) derivatives are organophosphates that are expected to reversibly bind within the BChE binding pocket in the same manner as natural physiological substrates. This study utilizes massive flexible inhibitor docking calculations to predict the relative binding affinities between the enzyme and a number of AACP derivatives and to predict the optimal binding orientation(s) of each AACP derivative within the BChE active site. When compared to their previously studied dialkyl phenyl phosphate (DAPP) analogs, AACP derivatives are predicted to interact more strongly with the binding site of BChE, thus serving as more potent inhibitors. Our results reproduce experimental trends in binding affinity, indicating that derivatives with longer alkyl chains show increased inhibitory strength (as observed for DAPP derivatives), and the neutralization of the charge of the choline group and protonation of the neutral choline group had little effect on inhibitor strength, offering insight into the structural preferences of BChE for specific AACP derivatives. This study thus provides a framework for both future inhibitor design and more rigorous molecular dynamics studies of cholinesterase-inhibitor interaction.

How to name atoms in phosphates, polyphosphates, their derivatives and mimics, and transition state analogues for enzyme-catalysed phosphoryl transfer reactions (IUPAC Recommendations 2016)

Procedures are proposed for the naming of individual atoms, P, O, F, N, and S in phosphate esters, amidates, thiophosphates, polyphosphates, their mimics, and analogues of transition states for enzyme-catalyzed phosphoryl transfer reactions. Their purpose is to enable scientists in very different fields, e.g. biochemistry, biophysics, chemistry, computational chemistry, crystallography, and molecular biology, to share standard protocols for the labelling of individual atoms in complex molecules. This will facilitate clear and unambiguous descriptions of structural results, as well as scientific intercommunication concerning them. At the present time, perusal of the Protein Data Bank (PDB) and other sources shows that there is a limited degree of commonality in nomenclature, but a large measure of irregularity in more complex structures. The recommendations described here adhere to established as closely as possible, in particular to IUPAC and IUBMB recommendations and to best practice in the PDB, especially to its atom labelling of amino acids, and particularly to Cahn-Ingold-Prelog rules for stereochemical nomenclature. They are designed to work in complex enzyme sites for binding phosphates but also to have utility for non-enzymatic systems. Above all, the recommendations are designed to be easy to comprehend and user-friendly.

A synthetic diphosphoinositol phosphate analogue of inositol trisphosphate.

Diphosphoinositol phosphates (PP-InsPs) are inositol phosphates (InsPs) that contain PP (diphosphate) groups. Converting a phosphate group in an InsP into a diphosphate has been reported to enhance affinity for some binding proteins. We synthesised 1-PP-Ins(4,5)P2, the first diphosphate analogue of the intracellular signalling molecule InsP3, and examined its effects on InsP3 receptors, which are intracellular Ca2+ channels. 1-PP-Ins(4,5)P2 was indistinguishable from InsP3 in its ability to bind to and activate type 1 InsP3 receptors, indicating that the diphosphate modification of InsP3 affected neither affinity nor efficacy. Nevertheless, 1-PP-Ins(4,5)P2 is the most potent 1-phosphate modified analogue of InsP3 yet identified. PP-InsPs are generally hydrolysed by diphosphoinositol phosphate phosphohydrolases (DIPPs), but 1-PP-Ins(4,5)P2 was not readily metabolised by human DIPPs. Differential scanning fluorimetry showed that 1-PP-Ins(4,5)P2 stabilises DIPP proteins, but to a lesser extent than naturally occurring substrates 1-PP-InsP5 and 5-PP-InsP5. The non-hydrolysable InsP7 analogues 1-PCP-InsP5 and 5-PCP-InsP5 showed comparable stabilising abilities to their natural counterparts and may therefore be promising substrate analogues for co-crystallisation with DIPPs.

Expression, purification, and characterization of a novel acid phosphatase that displays protein tyrosine phosphatases activity from Metarhizium anisopliae strain CQMa102.

The protein tyrosine phosphatase (PTPase) plays an important role in insect immune system. Our group has purified a type of acid phosphatase that could specifically dephosphorylate trans-Golgi p230 in vitro. In order to study this phosphatase further, we have identified and cloned the phosphatase gene from a locust specific Metarhizium anisopliae Strain CQMa102. The CQMa102 phosphatase was expressed in Pichia pastoris to verify its protease activity. The molecular weight (MW) and the isoelectric point (pI) of the phosphatase were about 85kDa and 6.15, respectively. Substrate specificity evaluation showed that the purified enzyme exhibited high activity on O-phospho-L-tyrosine. At its optimal pH of 6.5 and optimum temperature of 70°C, the protein showed the highest activity respectively. It can be activated by Ca2+, Mg2+, Mn2+, Ba2+, Co2+ and phosphate analogs, but inhibited by Zn2+, Cu2+, fluoride, dithiothreitol, β-mercaptoethanol and N-ethylmaleimide.

Tempo-phosphate as an ESR tool to study phosphate transport

TEMPO-phosphate has been introduced as a phosphate analogue to study phosphate transport in erythrocytes. The nitroxide is reduced intracellularly upon entering the cells, the membrane transport being the rate-limiting step of the loss of ESR signal. The use of TEMPO-phosphate is convenient and avoids the hazard of radioactivity. We studied the inhibition of TEMPO-phosphate transport to human erythrocytes by various compounds. DIDS and SITS, inhibitors of Band 3, inhibited the TEMPO-phosphate transport. 1-cyano-4-hydroxycinnamic acid, inhibitor of monocarboxylate transporters, did not affect the permeation of TEMPO-phosphate. The transport of TEMPO-phosphate was inhibited by various polyphenols, especially curcumin, naringin, quercetin, luteolin and kaempferol. Interestingly, 3-bromopyruvic acid, an alkylating agent and potential anticancer agent, induced an apparent enhancement of TEMPO-phosphate transport into erythrocytes.

Crystal structure of sodium di-hydrogen arsenate.

Single crystals of the title compound, Na(H2AsO4), were obtained by partial neutralization of arsenic acid with sodium hydroxide in aqueous solution. The crystal structure of Na(H2AsO4) is isotypic with the phosphate analogue and the asymmetric unit consists of two sodium cations and two tetra-hedral H2AsO4- anions. Each of the sodium cations is surrounded by six O atoms of five H2AsO4- groups, defining distorted octa-hedral coordination spheres. In the extended structure, the sodium cations and di-hydrogen arsenate anions are arranged in the form of layers lying parallel to (010). Strong hydrogen bonds [range of O⋯O distances 2.500 (3)-2.643 (3) Å] between adjacent H2AsO4- anions are observed within and perpendicular to the layers. The isotypic structure of Na(H2PO4) is comparatively discussed.

Metal cation controls phosphate release in the myosin ATPase.

Myosin is an enzyme that utilizes ATP to produce a conformational change generating a force. The kinetics of the myosin reverse recovery stroke depends on the metal cation complexed with ATP. The reverse recovery stroke is slow for MgATP and fast for MnATP. The metal ion coordinates the γ phosphate of ATP in the myosin active site. It is accepted that the reverse recovery stroke is correlated with the phosphate release; therefore, magnesium "holds" phosphate tighter than manganese. Magnesium and manganese are similar ions in terms of their chemical properties and the shell complexation; hence, we propose to use these ions to study the mechanism of the phosphate release. Analysis of octahedral complexes of magnesium and manganese show that the partial charge of magnesium is higher than that of manganese and the slightly larger size of manganese ion makes its ionic potential smaller. We hypothesize that electrostatics play a role in keeping and releasing the abstracted γ phosphate in the active site, and the stronger electric charge of magnesium ion holds γ phosphate tighter. We used stable myosin-nucleotide analog complex and Raman spectroscopy to examine the effect of the metal cation on the relative position of γ phosphate analog in the active site. We found that in the manganese complex, the γ phosphate analog is 0.01 nm further away from ADP than in the magnesium complex. We conclude that the ionic potential of the metal cation plays a role in the retention of the abstracted phosphate.

The interactions of metal cations and oxyanions with protein tyrosine phosphatase 1B

Protein tyrosine phosphatases are not considered to be metalloenzymes. Yet, they are inhibited by zinc cations and metal and non-metal oxyanions that are chemical analogues of phosphate, e.g. vanadate. Metal inhibition is generally not recognized as these enzymes are purified, supplied, and assayed with buffers containing chelating and reducing agents. We screened a series of cations and anions for their capacity to inhibit protein tyrosine phosphatase 1B and discuss the ensuing general issues with inhibition constants reported in the scientific literature. In contrast to zinc, which binds to the phosphocysteine intermediate in the closed conformation of protein tyrosine phosphatase 1B when the catalytic aspartate has moved into the active site, other divalent cations such as cadmium and copper may also bind to the enzyme in the open conformation. Inhibition by both anions and cations, conditions such as pH, the presence of metal ligands such as glutathione, and the existence of multiple conformational states of protein tyrosine phosphatases in the reaction cycle establish a complex pattern of inhibition of these important regulatory enzymes with implications for the physiology, pharmacology and toxicology of metal ions.

Identification of metabolically stable 5΄-phosphate analogs that support single-stranded siRNA activity.

Identification of metabolically stable 5΄-phosphate analogs that support single-stranded siRNA activity.

How to name atoms in phosphates, polyphosphates, their derivatives and mimics, and transition state analogues for enzyme-catalysed phosphoryl transfer reactions (IUPAC Recommendations 2016)

Abstract Procedures are proposed for the naming of individual atoms, P, O, F, N, and S in phosphate esters, amidates, thiophosphates, polyphosphates, their mimics, and analogues of transition states for enzyme-catalyzed phosphoryl transfer reactions. Their purpose is to enable scientists in very different fields, e.g. biochemistry, biophysics, chemistry, computational chemistry, crystallography, and molecular biology, to share standard protocols for the labelling of individual atoms in complex molecules. This will facilitate clear and unambiguous descriptions of structural results, as well as scientific intercommunication concerning them. At the present time, perusal of the Protein Data Bank (PDB) and other sources shows that there is a limited degree of commonality in nomenclature, but a large measure of irregularity in more complex structures. The recommendations described here adhere to established practice as closely as possible, in particular to IUPAC and IUBMB recommendations and to “best practice” in the PDB, especially to its atom labelling of amino acids, and particularly to Cahn-Ingold-Prelog rules for stereochemical nomenclature. They are designed to work in complex enzyme sites for binding phosphates but also to have utility for non-enzymatic systems. Above all, the recommendations are designed to be easy to comprehend and user-friendly.

A catalytic approach to synthesis of PLP analogs and other environmental protocols in a single handed CaO/TiO2 green nanoparticle

As our precursory stage we have focus straight forward on clean catalytic approach for the production of C3 substituted pyridoxal-5′-phosphate analogues of vitamin B6, and other environmental protocols like photocatalytic activity, green fossil fuels and c–c coupling using efficient biocompatible eggshell related unrivalled materials which show versatility of the catalytic effect on different inorganic support. The eggshell immobilized nanoparticles have encouraging relevance in creation of new molecules and can advantageously be studied by various spectroscopic, thermal and elemental analyses like powder X-ray diffraction (XRD), Raman spectroscopy, UV–vis, Scanning electron microscopy (SEM), Energy dispersive X-ray analysis (EDX), X-ray photoelectron spectroscopy (XPS) and Brunauer-Emmett-Teller (BET) surface area analysis. The elucidate nature of nanoparticles offer: more active site acts as lewis acid, vacancies on the catalyst surface and good to better yield of C3 substituted deoxy and 2-nor deoxy coenzyme pyridoxine (PN), coupling products propargylamines (PA), photo degrading enhancement of MB and nucleophilic substituted fatty acid (BD). This enzyme cofactor explore molecular synthons to synthetic equivalent: 3-deoxy and 2-nor-3-deoxy pyridoxal (PL), pyridoxal oxime (PO), pyridoxamine (PM) and mono phosphate derivative of 3-deoxyPM, 3-deoxyPL respectively and chemistry of selective oxidation and schiff base mechanism was studied and complemented through combined experimental and theoretical molecular orbital calculation consequently. The heterogeneous catalyst has strong selective ability towards selective reducing pyridine diester, bioactive intermediates substances and holds vast potential towards separation for the photogenerated electron-hole pairs and renewable, nontoxic, biodegradable green fossil fuels. The catalyst including environmental concern is reapplicable and strong impressive that can unfold the space of worthy metal component widely and facilitate the scope to take a vital role in different fileds like catalysis, biochemistry, nanoscience, energy and materials science.

Ability of Periplasmic Phosphate Binding Proteins from Synechocystis sp. PCC 6803 to Discriminate Phosphate Against Arsenate

Arsenate (As(V)) is an analog of phosphate (Pi), and previous studies have shown that As(V) and Pi are absorbed via the same transport systems in some organisms. However, little is known about which periplasmic phosphate-binding proteins (PBPs) of ABC-type Pi transporters play major roles in As(V) uptake by cyanobacteria. In this study, the model cyanobacterium Synechocystis sp. PCC 6803 (Synechocystis) was chosen to study the ability of PBPs to discriminate between Pi and As(V). We found that As(V) and Pi competed for uptake via Pi transporters when Synechocystis was treated with As(V) and/or Pi in short-term incubation. The As/P molar ratios of Synechocystis wild type (WT) and mutants (∆sphX, ∆pstS2, ∆sll0540, and ∆sphXpstS1) were measured, and they were significantly different with the order WT > ∆pstS2 > ∆sll0540 > ∆sphXpstS1 > ∆sphX. Furthermore, As(V) uptake by Escherichia coli strain Transetta expressing PBP genes, particularly sphX or sll0540, was significantly increased when PBP gene (pstS1, pstS2, sphX, or sll0540) was separately expressed in Transetta at the same level. Subsequent phylogenetic analyses of PBPs showed that SphX and Sll0540 belonged to the low-affinity PBPs, while PstS1 and PstS2 were clustered with the high-affinity PBPs. These results implied that As(V) was taken up via Pi transporters, and the low-affinity PBPs, SphX and Sll0540, made a significant contribution to As(V) uptake.

Phosphorus deficiency modifies As translocation in the halophyte plant species Atriplex atacamensis

Most arsenic in surface soil and water exists primarily in its oxidized form, as arsenate (As(V); AsO43-), which is an analog of phosphate (PO43-). Arsenate can be taken up by phosphate transporters. Atriplex atacamensis Phil. is native to northern Chile (Atacama Desert), and this species can cope with high As concentrations and low P availability in its natural environment. To determine the impact of P on As accumulation and tolerance in A. atacamensis, the plants were cultivated in a hydroponic system under four treatments: no As(V) addition with 323µM phosphate (control); 1000µM As(V) addition with 323µM phosphate; no As(V) and no phosphate; 1000µM As(V) addition and no phosphate. Phosphate starvation decreased shoot fresh weight, while As(V) addition reduced stem and root fresh weights. Arsenate addition decreased the P concentrations in both roots and leaves, but to a lesser extent than for P starvation. Phosphorus starvation increased the As concentrations in roots, but decreased it in shoots, which suggests that P deficiency reduced As translocation from roots to shoots. Arsenate addition increased total glutathione, but P deficiency decreased oxidized and reduced glutathione in As(V)-treated plants. Arsenate also induced an increase in S accumulation and nonprotein thiol and ethylene synthesis, and a decrease in K concentrations, effects that were similar for the P-supplied and P-starved plants. In contrast, in As(V)-treated plants, P starvation dramatically decreased total soluble protein content and increased lipid peroxidation, compared to plants supplied with P. Phosphorus nutrition thus appears to be an important component of A. atacamensis response to As toxicity.

Crystal structure and biochemical characterization of the transmembrane PAP2 type phosphatidylglycerol phosphate phosphatase from Bacillus subtilis.

Type 2 phosphatidic acid phosphatases (PAP2s) can be either soluble or integral membrane enzymes. In bacteria, integral membrane PAP2s play major roles in the metabolisms of glycerophospholipids, undecaprenyl-phosphate (C55-P) lipid carrier and lipopolysaccharides. By in vivo functional experiments and biochemical characterization we show that the membrane PAP2 coded by the Bacillus subtilis yodM gene is the principal phosphatidylglycerol phosphate (PGP) phosphatase of B. subtilis. We also confirm that this enzyme, renamed bsPgpB, has a weaker activity on C55-PP. Moreover, we solved the crystal structure of bsPgpB at 2.25Å resolution, with tungstate (a phosphate analog) in the active site. The structure reveals two lipid chains in the active site vicinity, allowing for PGP substrate modeling and molecular dynamic simulation. Site-directed mutagenesis confirmed the residues important for substrate specificity, providing a basis for predicting the lipids preferentially dephosphorylated by membrane PAP2s.