Phosphate Analogues Research Articles

Global profile changes in transcripts induced with a phosphate analogue: implications for gene regulation.

Previous work has shown that thiophosphate, a phosphate analogue, leads to a global shift in the distribution of cellular proteins in a variety of organisms. Thiophosphate, when added to culture media, gets incorporated into the nucleic acids of cells resulting in nuclease-resistant phosphorothioate linkages. Using Escherichia coli, as a model organism, it was found that the global changes in protein expression induced with thiophosphate could be accounted for by significant changes in the absolute transcription levels of more than 1500 genes detected via RNA seq analysis. In fact, 58% of transcripts detected in RNA seq studies using total RNA were increased an average of 44 × fold while the remaining 42% were decreased an average of 20 × fold in thiophosphate-treated cells. Furthermore, microarray results showed no correlation between the transcriptional changes observed and the known stability of the corresponding mRNAs measured. Overall, the total amount of non-ribosomal RNA accumulated in TP-treated cells was increased relative to rRNA ~ 4 × fold (1.5-6 ×). The results further indicated that metabolic changes may play a role in inducing the transcriptional profiles observed with thiophosphate. Indeed, pathway analysis of transcripts showed an increase in routes for phosphoribosyl pyrophosphate (PRPP) synthesis and related derivatives, presumably due to a reduction in RNA turnover. These results raise the possibility that the energy savings with reduced RNA turnover could lead to an increased energy charge in the cell that induces transcriptional changes leading to an increase in biosynthetic processes.

Inhibition of vascular calcification by inositol phosphates derivatized with ethylene glycol oligomers

Myo-inositol hexakisphosphate (IP6) is a natural product known to inhibit vascular calcification (VC), but with limited potency and low plasma exposure following bolus administration. Here we report the design of a series of inositol phosphate analogs as crystallization inhibitors, among which 4,6-di-O-(methoxy-diethyleneglycol)-myo-inositol-1,2,3,5-tetrakis(phosphate), (OEG2)2-IP4, displays increased in vitro activity, as well as more favorable pharmacokinetic and safety profiles than IP6 after subcutaneous injection. (OEG2)2-IP4 potently stabilizes calciprotein particle (CPP) growth, consistently demonstrates low micromolar activity in different in vitro models of VC (i.e., human serum, primary cell cultures, and tissue explants), and largely abolishes the development of VC in rodent models, while not causing toxicity related to serum calcium chelation. The data suggest a mechanism of action independent of the etiology of VC, whereby (OEG2)2-IP4 disrupts the nucleation and growth of pathological calcification.

Mitochondrial alternative oxidase (AOX1a) is required for the mitigation of arsenic-induced oxidative stress in Arabidopsis thaliana

The element arsenic (As) is a non-essential metalloid that is found naturally in all soils and at high concentrations it is toxic to plant cells. As (V) can act as a chemical analogue of phosphate, it can disrupt phosphate-related energy metabolism and lipid structure. In this study, the contribution of mitochondrial alternative oxidase (AOX) and chloroplastic plastid terminal oxidase (PTOX) to As (V) stress tolerance was investigated. Our data indicate that As (V) stress (100, 200 and 300 µM) induces AOX gene expression by 3.3- to 10.5-fold depending on AOX gene, but not PTOX expression in wild-type A. thaliana plants. To further elucidate the role of AOX in As (V) stress tolerance, we utilized aox1a mutants and observed that aox1a mutants had decreased growth and higher oxidative stress damage under stress conditions, while there were no differences under control conditions. Moreover, acclimation of aox1a plants to new cellular redox environment was investigated by measuring the activities of reactive oxygen species (ROS)-scavenging enzymes. Induction of mitochondrial MnSOD activity at 300 µM As (V) was higher in aox1a plants (70%), when compared to wild type (43%). However, total ascorbate peroxidase and dehydroascorbate reductase activities were lower in aox1a plants when compared to wild type, which might explain higher oxidative damage observed in this genotype. On the other hand, NADPH oxidase activity, which is involved in ROS signaling, was lower in aox1a plants under normal conditions but a higher induction was observed with As (V) stress. Overall, our data indicate that AOX1a is involved in adaptation to As (V)-induced oxidative stress.

Exogenous phosphite application alleviates the adverse effects of heat stress and improves thermotolerance of potato (Solanum tuberosum L.) seedlings.

Phosphite (Phi), an analog of phosphate (Pi) anion, is emerging as a potential biostimulator, fungicide and insecticide. Here, we reported that Phi also significantly enhanced thermotolerance in potatoes under heat stress. Potato plants with and without Phi pretreatment were exposed to heat stress and their heat tolerance was examined by assessing the morphological characteristics, photosynthetic pigment content, photosystem II (PS II) efficiency, levels of oxidative stress, and level of DNA damage. In addition, RNA-sequencing (RNA-Seq) was adopted to investigate the roles of Phi signals and the underlying heat resistance mechanism. RNA-Seq revealed that Phi orchestrated plant immune responses against heat stress by reprograming global gene expressions. Results from physiological data combined with RNA-Seq suggested that the supply of Phi not only was essential for the better plant performance, but also improved thermotolerance of the plants by alleviating oxidative stress and DNA damage, and improved biosynthesis of osmolytes and defense metabolites when exposed to unfavorable thermal conditions. This is the first study to explore the role of Phi in thermotolerance in plants, and the work can be applied to other crops under the challenging environment.

Antagonistic co-limitation through ion promiscuity – On the metal sensitivity of Thalassiosira oceanica under phosphorus stress

Nutrient limitation of primary producers is a fundamental principle in biogeochemical oceanography and has been used with great success in prescribing understanding to patterns of marine primary productivity. In recent years the paradigm of nutrient limitation has expanded from single nutrient limitation towards concepts of co-limitation by multiple resources. Interactive effects between multiple limiting resources are now thought commonplace in marine microbial communities. Here we investigate the response exhibited by phosphate-limited Thalassiosira oceanica to elevated concentrations of the phosphate analogs vanadate, arsenate and molybdate. Enrichments in external arsenate and vanadate to phosphate-limited cultures act to suppress growth rates entirely, an effect not seen in phosphate replete conditions. Retardation of growth rates is attributed to mistaken uptake through ion promiscuity as evidenced by observations of significant intracellular accumulation of both arsenic and vanadium under phosphate limited conditions. We describe this novel co-limitation scenario as dependent antagonistic co-limitation (DAC), and suggest that this phenomenon of non-deliberate intracellular accumulation could be used as both a proxy of phosphate stress in the modern ocean and a possible marker of phosphate depletion limiting the duration of oceanic anoxic events.

SPX4 Acts on PHR1-Dependent and -Independent Regulation of Shoot Phosphorus Status in Arabidopsis.

Phosphorus (P) is an essential macronutrient for all living organisms and limits plant growth. Four proteins comprising a single SYG1/Pho81/XPR1 (SPX) domain, SPX1 to SPX4, are putative phosphate-dependent inhibitors of Arabidopsis (Arabidopsis thaliana) PHOSPHATE STARVATION RESPONSE1 (PHR1), the master transcriptional activator of phosphate starvation responses. This work demonstrated that SPX4 functions as a negative regulator not only of PHR1-dependent but also of PHR1-independent responses in P-replete plants. Transcriptomes of P-limited spx4 revealed that, unlike SPX1 and SPX2, SPX4 modulates the shoot phosphate starvation response but not short-term recovery after phosphate resupply. In roots, transcriptional regulation of P status is SPX4 independent. Genes misregulated in spx4 shoots intersect with both PHR1-dependent and PHOSPHATE2-dependent signaling networks associated with plant development, senescence, and ion/metabolite transport. Gene regulatory network analyses suggested that SPX4 interacts with transcription factors other than PHR1, such as SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 and ARABIDOPSIS NAC DOMAIN CONTAINING PROTEIN55, known regulators of shoot development. Transient expression studies in protoplasts indicated that PHR1 retention in the cytosol by SPX4 occurs in a dose- and P-status-dependent manner. Using a luciferase reporter in vivo, SPX4 expression kinetics and stability revealed that SPX4 is a short-lived protein with P-status-dependent turnover. SPX4 protein levels were quickly restored by phosphate resupply to P-limited plants. Unlike its monocot ortholog, AtSPX4 was not stabilized by the phosphate analog phosphite, implying that intracellular P status is sensed by its SPX domain via phosphate-rich metabolite signals.

Levantite, KCa3(Al2Si3)O11(PO4), a new latiumite-group mineral from the pyrometamorphic rocks of the Hatrurim Basin, Negev Desert, Israel

AbstractLevantite, with the end-member formula KCa3(Al2Si3)O11(PO4), is the phosphate analogue of latiumite, KCa3(Al3Si2)O11(SO4, CO3) found in gehlenite–wollastonite hornfels on Har Parsa, Negev Desert, Israel. Levantite forms later zones on long-prismatic crystals of latiumite. Rarer homogeneous colourless levantite crystals up to 0.2 mm long and with mean composition (K0.94Ba0.01Na0.01□0.04)Σ1.00(Ca2.96Mg0.03)Σ2.99{(Si2.69Al2.06Fe3+0.16P0.06)Σ4.97O11}[(PO4)0.65(SO4)0.35]Σ1.00were noted. Minerals of the levantite–latiumite series are associated with gehlenite, wollastonite, clinopyroxene of the esseneite–diopside series, anorthite and Ti-bearing andradite. Levantite crystalises in space groupP21with unit-cell parametersa= 12.1006(9) Å,b= 5.1103(4) Å,c= 10.8252(9) Å, β = 107.237(8)°,V= 639.34(9) Å3andZ= 2. The structure of levantite is analogous to latiumite. It is formed by tetrahedral hybridzweierdouble layers [(Si,Al)10O22] connected by Ca atoms. Three Ca atoms linked to different double layers are bridged over by (PO4) and minor (SO4) groups. K atoms reside in the cavities between two superimposedzweierdouble layers. The measured micro-indentation hardness of levantite gave VHN50= 580(19) (mean of 14), range 550–611 kg/mm2, which correlates with 5 on the Mohs scale. Cleavage is good on (100). Twinning on (100) is polysynthetic or simple. The calculated density is 2.957 g cm–3. Levantite is optically negative with α = 1.608(2), β = 1.618(2), γ = 1.622(2) (λ = 589 nm), 2Vmeas.= 70(5)° and 2Vcalc.= 64.3°. Dispersion of the optical axes r > v is weak; the optical orientation is:Z=b,X^c= 22–27°; and it is non-pleochroic. Minerals of the levantite–latiumite series from Israel show characteristic Raman spectra with the main bands at 994 cm–1[ν1(SO4)2–] and 945 cm–1[ν1(PO4)3–]. The band intensity ν1(PO4)3–/ν1(SO4) ratio is well correlated with P and S contents in the investigated minerals. The strongest lines in the powder diffraction pattern [dobs, Å (I, %) (hkl)] are: 3.0762(100)(310), 2.8551(96)($\bar 2$13), 2.9704(92)($\bar 3$12), 2.8573(83)(013), 2.5552(66)(020), 2.8228(48)(212), 2.8893(40)(400), and 3.0634(30)(103).

Instructed-Assembly of Small Peptides Inhibits Drug-Resistant Prostate Cancer Cells.

Despite multiple new-drug approvals in recent years, prostate cancer remains a global health challenge because of the prostate cancers are resistant to androgen deprivation therapy. Here we show that a small D-phosphopeptide undergoes prostatic acid phosphatase (PAP)-instructed self-assembly for inhibiting castration-resistant prostate cancer (CRPC) cells. Specifically, the installation of phosphate at the C-terminal of a D-tripeptide results in the D-phosphopeptide. Dephosphorylating the D-phosphopeptide by PAP forms uniform nanofibers that inhibit VCaP, a castration-resistant prostate cancer cell. A non-hydrolyzable phosphate analogue of the D-phosphopeptide, which shares similar self-assembling properties with the D-phosphopeptide, confirms that PAP-instructed assembly is critical for the inhibition of VCaP. This work, for the first time, demonstrates PAP-instructed self-assembly of peptides for selective inhibiting castration-resistant prostate cancer (CRPC) cells.

Geochemical Sources and Availability of Amidophosphates on the Early Earth

AbstractPhosphorylation of (pre)biotically relevant molecules in aqueous medium has recently been demonstrated using water‐soluble diamidophosphate (DAP). Questions arise relating to the prebiotic availability of DAP and other amidophosphosphorus species on the early earth. Herein, we demonstrate that DAP and other amino‐derivatives of phosphates/phosphite are generated when Fe3P (proxy for mineral schreibersite), condensed phosphates, and reduced oxidation state phosphorus compounds, which could have been available on early earth, are exposed to aqueous ammonia solutions. DAP is shown to remain in aqueous solution under conditions where phosphate is precipitated out by divalent metals. These results show that nitrogenated analogues of phosphate and reduced phosphite species can be produced and remain in solution, overcoming the thermodynamic barrier for phosphorylation in water, increasing the possibility that abiotic phosphorylation reactions occurred in aqueous environments on early earth.

Geochemical Sources and Availability of Amidophosphates on the Early Earth.

Phosphorylation of (pre)biotically relevant molecules in aqueous medium has recently been demonstrated using water-soluble diamidophosphate (DAP). Questions arise relating to the prebiotic availability of DAP and other amidophosphosphorus species on the early earth. Herein, we demonstrate that DAP and other amino-derivatives of phosphates/phosphite are generated when Fe3 P (proxy for mineral schreibersite), condensed phosphates, and reduced oxidation state phosphorus compounds, which could have been available on early earth, are exposed to aqueous ammonia solutions. DAP is shown to remain in aqueous solution under conditions where phosphate is precipitated out by divalent metals. These results show that nitrogenated analogues of phosphate and reduced phosphite species can be produced and remain in solution, overcoming the thermodynamic barrier for phosphorylation in water, increasing the possibility that abiotic phosphorylation reactions occurred in aqueous environments on early earth.

(Cs X)Cu5O2(PO4)2 ( X = Cl, Br, I): A Family of Cu2+ S = 1/2 Compounds with Capped-Kagomé Networks Composed of OCu4 Units.

Three new salt inclusion compounds (Cs X)Cu5O2(PO4)2 ( X = Cl, Br, I), phosphate analogues of the kagomé mineral averievite, are reported. Their crystal structures are composed of trigonal networks of corner-sharing OCu4 anion-centered tetrahedra, forming capped-kagomé planes, which can also be regarded as two-dimensional slices along the [111] direction of a pyrochlore lattice. Magnetization and heat capacity measurements reveal strong geometric frustration of this network and complex magnetic behavior. X-ray and neutron diffraction studies show that all three compounds undergo a trigonal-to-monoclinic phase transition upon cooling, with a first-order phase transition seen in CsBr and CsI analogues. Along with the previously reported (CsCl)Cu5O2(VO4)2, these three new compounds belong to a large family of OCu4-based networks, which are a playground for studying frustrated quantum magnetism.

Characterization of phoA, a Bacterial Alkaline Phosphatase for Phi Use Efficiency in Rice Plant.

Fertilizers and herbicides are two major components in the agriculture system for achieving crop productivity. Massive use of orthophosphate fertilizers and herbicides poses threats to phosphate reserves and aids the evolution of herbicide tolerant weed biotypes. Phosphite (Phi), a phosphate analog, has been proposed as more beneficial than traditionally used phosphate fertilizers and herbicides in the agriculture. We developed phoA overexpressing transgenic rice that minimizes the phosphate loss and contributes to weed management in the agriculture. The phoA rice lines showed improved root, shoot length and total biomass production under phosphite conditions. Additionally, the complete phenotype and productivity of phoA lines under the phosphite treatment attained was similar to that of plants under phosphate sufficient condition. The Phi metabolizing properties of the phoA overexpressed lines improved under the Phi application and phi treatment enabled controlling of weeds without compromising the yield of transgenic rice plants. Our results indicated that phoA alone or in combination with other Phi metabolizing gene(s) can possibly be used as an effective ameliorating system for improving crop plants for phi-based fertilization and weed management strategy in the agriculture.

CdSe/ZnS quantum dots coated with carboxy-PEG and modified with the terbium(III) complex of guanosine 5'-monophosphate as a fluorescent nanoprobe for ratiometric determination of arsenate via its inhibition of acid phosphatase activity.

A ratiometric fluorometric method is described for the determination of arsenate via its inhibitory effect on the activity of the enzyme acid phosphatase. A nanoprobe was designed that consists of CdSe/ZnS quantum dots (QDs) coated with the terbium(III) complex of guanosine monophosphate (Tb-GMP). The nanoprobe was synthesized from carboxylated QDs, Tb(III) and GMP via binding of Tb(III) by both the carboxy and the phosphate groups. The nanoprobe, under single-wavelength excitation (at 280nm), displays dual (red and green) emission, with peaks at around 652nm from the QDs, and at 547nm from the Tb-GMP coordination polymers. It is shown to be a viable nanoprobe for fluorometric determination of As(V) detection through it inhibitory action on the activity of acid phosphatase (ACP). The enzyme destroys the Tb-GMP structure via hydrolysis of GMP, and hence the fluorescence of the Tb-GMP complex is quenched. In contrast, the fluorescence of the CdSe/ZnS QDs remains inert to ACP. It therefore can serve as an internal reference signal. In the presence of arsenate (an analog of phosphate), the activity of ACP is inhibited due to competitive binding. Thus, hydrolysis of GMP is prevented. These findings were used to design a ratiometric fluorometric method for the quantification of As(V). The ratio of fluorescences at 547 and 652nm increases linearly in the 0.5 to 200ppb As(V) concentation range, and the limit of detection is 0.39ppb. Under a UV lamp, the probe shows distinguishable color from green to red on increasing the concentration of As(V). Graphical abstract Schematic illustration of CdSe/ZnS quantum dot coated with carboxy-PEG and modified with the terbium(III)-GMP complex as a fluorescent nanoprobe for ratiometric determination of arsenate via its inhibition of ACP activity.

Interaction landscape of a 'CαNN' motif with arsenate and arsenite: a potential peptide-based scavenger of arsenic.

Arsenic (As) is a toxic metalloid that has drawn immense attention from the scientific community recently due to its fatal effects through its unwanted occurrence in ground water around the globe. The presence of an excess amount of water soluble arsenate and/or arsenite salt (permissible limit 10 μg L−1 as recommended by the WHO) in water has been correlated with several human diseases. Although arsenate (HAsO42−) is a molecular analogue of phosphate (HPO42−), phosphate is indispensable for life, while arsenic and its salts are toxic. Therefore, it is worthwhile to focus on the removal of arsenic from water. Towards this end, the design of peptide-based scaffolds for the recognition of arsenate and arsenite would add a new dimension. Utilizing the stereochemical similarity between arsenate (HAsO42−) and phosphate (HPO42−), we successfully investigated the recognition of arsenate and arsenite with a naturally occurring novel phosphate binding ‘CαNN’ motif and its related designed analogues. Using computational as well as biophysical approaches, for the first time, we report here that a designed peptide-based scaffold based on the ‘CαNN’ motif can recognize anions of arsenic in a thermodynamically favorable manner in a context-free system. This peptide-based arsenic binding agent has the potential for future development as a scavenger of arsenic anions to obtain arsenic free water.

Biostimulant and fungicidal effects of phosphite assessed by GC-TOF-MS analysis of potato leaf metabolome

Phosphite (Phi), an analog of phosphate, is increasingly used worldwide as fungicide, fertilizer and plant biostimulant. Studies using various plant systems provided insights on how Phi is translocated in plants and on its mode of action against pathogens. However, little evidence is present at the metabolite level, of the compounds present in plants that are responsible in the enhanced resistance to pathogens. As Phi is used to control late blight, the most devastating disease of potato, caused by the oomycete Phytophthora infestans, this study used potato as a model plant to study the effects of this chemical. Analysis of potato leaf metabolome, using GC-TOF-MS, revealed that several metabolite pools are modified by Phi applications. Phi treatments were found to be associated with higher levels of components of the phenylpropanoid pathway such as chlorogenic acid, caffeic acid and salicylic acid, compounds that are known to be essential components of plant defense response to biotic and abiotic stresses, providing therefore additional support to an effective indirect mode of action.

Synthesis and Kinetic evaluation of an azido analogue of methylerythritol phosphate: a Novel Inhibitor of E. coli YgbP/IspD

As multidrug resistant pathogenic microorganisms are a serious health menace, it is crucial to continuously develop novel medicines in order to overcome the emerging resistance. The methylerythritol phosphate pathway (MEP) is an ideal target for antimicrobial development as it is absent in humans but present in most bacteria and in the parasite Plasmodium falciparum. Here, we report the synthesis and the steady-state kinetics of a novel potent inhibitor (MEPN3) of Escherichia coli YgbP/IspD, the third enzyme of the MEP pathway. MEPN3 inhibits E. coli YgbP/IspD in mixed type mode regarding both substrates. Interestingly, MEPN3 shows the highest inhibitory activity when compared to known inhibitors of E. coli YgbP/IspD. The mechanism of this enzyme was also studied by steady-state kinetic analysis and it was found that the substrates add to the enzyme in sequential manner.

E2P-like states of plasma membrane Ca2+‑ATPase characterization of vanadate and fluoride-stabilized phosphoenzyme analogues

The plasma membrane Ca2+‑ATPase (PMCA) belongs to the family of P-type ATPases, which share the formation of an acid-stable phosphorylated intermediate as part of their reaction cycle. The crystal structure of PMCA is currently lacking. Its abundance is approximately 0.1% of the total protein in the membrane, hampering efforts to produce suitable crystals for X-ray structure analysis. In this work we characterized the effect of beryllium fluoride (BeFx), aluminium fluoride (AlFx) and magnesium fluoride (MgFx) on PMCA. These compounds are known inhibitors of P-type ATPases that stabilize E2P ground, E2·P phosphoryl transition and E2·Pi product states. Our results show that the phosphate analogues BeFx, AlFx and MgFx inhibit PMCA Ca2+‑ATPase activity, phosphatase activity and phosphorylation with high apparent affinity. Ca2+‑ATPase inhibition by AlFx and BeFx depended on Mg2+ concentration indicating that this ion stabilizes the complex between these inhibitors and the enzyme. Low pH increases AlFx and BeFx but not MgFx apparent affinity. Eosin fluorescent probe binds with high affinity to the nucleotide binding site of PMCA. The fluorescence of eosin decreases when fluoride complexes bind to PMCA indicating that the environment of the nucleotide binding site is less hydrophobic in E2P-like states. Finally, measuring the time course of E → E2P-like conformational change, we proposed a kinetic model for the binding of fluoride complexes and vanadate to PMCA.In summary, our results show that these fluoride complexes reveal different states of phosphorylated intermediates belonging to the mechanism of hydrolysis of ATP by the PMCA.

A kinetic comparison between E2P and the E2P-like state induced by a beryllium fluoride complex in the Na,K-ATPase. Interactions with Rb+

Metal-fluoride complexes have been used to induce E2P-like states with the aim of studying the events that occur during E2P hydrolysis in P-type ATPases. In the present work, we compared the E2P-like state induced by a beryllium fluoride complex (BeFx) with the actual E2P state formed through backdoor phosphorylation of the Na,K-ATPase. Formation of E2P and E2P-like states were investigated employing the styryl dye RH421. We found that BeFx is the only fluorinated phosphate analog that, like Pi, increases the RH421 fluorescence. The observed rate constant, kobs, for the formation of E2P decreases with [Pi] whereas that of E2BeFx increases with [BeFx]. This might wrongly be taken as evidence of a mechanism where the binding of BeFx induces a conformational transition. Here, we rather propose that, like for Pi, binding of BeFx follows a conformational-selection mechanism, i.e. it binds to the E2 conformer forming a complex that is much more stable than E2P, as seen from its impaired capacity to return to E1 upon addition of Na+. Although E2P and E2BeFx are able to form states with 2 occluded Rb+, both enzyme complexes differ in that the affinity for the binding and occlusion of the second Rb+ is much lower in E2BeFx than in E2P. The higher rates of Rb+ occlusion and deocclusion observed for E2BeFx, as compared to those observed for other E2P-like transition and product states suggest a more open access to the cation transport sites, supporting the idea that E2BeFx mimics the E2P ground state.

Dissection of differential vanadate sensitivity in two Ogataea species links protein glycosylation and phosphate transport regulation

The closely related yeasts Ogataea polymorpha and O. parapolymorpha differ drastically from each other by sensitivity to the toxic phosphate analog vanadate. Search for genes underlying this difference revealed two genes, one designated as ABV1 (Alcian Blue staining, Vanadate resistance), which encodes a homologue of Saccharomyces cerevisiae Mnn4 responsible for attachment of mannosylphosphate to glycoside chains of secretory proteins, and the other designated as its S. cerevisiae homologue PHO87, encoding the plasma membrane low affinity phosphate sensor/transporter. The effect of Pho87 on vanadate resistance was bidirectional, since it decreased the resistance on phosphate-depleted medium, but was required for pronounced protection against vanadate by external phosphate. This highlights the dual function of this protein as a low affinity phosphate transporter and an external phosphate sensor. Involvement of Pho87 in phosphate sensing was confirmed by its effects on regulation of the promoter of the PHO84 gene, encoding a high affinity phosphate transporter. The effect of Abv1 was also complex, since it influenced Pho87 level and enhanced repression of the PHO84 promoter via a Pho87-independent pathway. Role of the identified genes in the difference in vanadate resistance between O. polymorpha and O. parapolymorpha is discussed.

Effect of arsenate substitution on phosphate repository of cell: a computational study.

The structural analogy with phosphate derives arsenate into various metabolic processes associated with phosphate inside the organisms. But it is difficult to evaluate the effect of arsenate substitution on the stability of individual biological phosphate species, which span from a simpler monoester form like pyrophosphate to a more complex phosphodiester variant like DNA. In this study, we have classified the physiological phosphate esters into three different classes on the basis of their structural differences. This classification has helped us to present a concise theoretical study on the kinetic stability of phosphate analogue species of arsenate against hydrolysis. All the calculations have been carried out using QM/MM methods of our Own N-layer Integrated molecular Orbital molecular Mechanics (ONIOM). For quantum mechanical region, we have used M06-2X density functional with 6-31+G(2d,2p) basis set and for molecular mechanics we have used the AMBER force field. The calculated rate constants for hydrolysis show that none of the phosphate analogue species of arsenate has a reasonable stability against hydrolysis.