Foreword to the First Edition.Preface.Acknowledgements.Introduction.1 The Chemical Structures and Properties of Condensed Inorganic Phosphates.1.1 The Structures of Condensed Phosphates.1.1.1 Cyclophosphates.1.1.2 Polyphosphates.1.1.3 Branched Inorganic Phosphates, or 'Ultraphosphates'.1.2 Some Chemical Properties of Condensed Inorganic Polyphosphates.1.3 Physico-Chemical Properties of Condensed Inorganic Polyphosphates.2 Methods of Polyphosphate Assay in Biological Materials.2.1 Methods of Extraction from Biological Materials.2.2 Chromatographic Methods.2.3 Colorimetric and Fluorimetric Methods.2.4 Cytochemical Methods.2.5 X-Ray Energy Dispersive Analysis.2.6 31P Nuclear Magnetic Resonance Spectroscopy.2.7 Other Physical Methods.2.8 Gel Electrophoresis.2.9 Enzymatic Methods.3 The Occurrence of Polyphosphates in Living Organisms.4 The Forms in which Polyphosphates are Present in Cells.4.1 Polyphosphate-Cation Complexes.4.2 Polyphosphate-Ca2+-Polyhydroxybutyrate Complexes.4.3 Complexes of Polyphosphates with Nucleic Acids.4.4 Binding of Polyphosphates with Proteins.5 Localization of Polyphosphates in Cells of Prokaryotes and Eukaryotes.5.1 Prokaryotes.5.2 Eukaryotes.6 Enzymes of Polyphosphate Biosynthesis and Degradation.6.1 Enzymes of Polyphosphate Biosynthesis.6.1.1 Polyphosphate Kinase (Polyphosphate:ADP Phosphotransferase, EC 2.7.4.1).6.1.2 3-Phospho-D-Glyceroyl-Phosphate:Polyphosphate Phosphotransferase (EC 2.7.4.17).6.1.3 Dolichyl-Diphosphate:Polyphosphate Phosphotransferase (EC 2.7.4.20).6.2 Enzymes of Polyphosphate Degradation.6.2.1 Polyphosphate-Glucose Phosphotransferase (EC 2.7.1.63).6.2.2 NAD Kinase (ATP:NAD 2--Phosphotransferase, EC 2.7.1.23).6.2.3 Exopolyphosphatase (Polyphosphate Phosphohydrolase, EC 3.6.1.11).6.2.4 Adenosine-Tetraphosphate Phosphohydrolase (EC 3.6.1.14).6.2.5 Triphosphatase (Tripolyphosphatase, EC 3.6.1.25).6.2.6 Endopolyphosphatase (Polyphosphate Depolymerase, EC 3.6.1.10).6.2.7 PolyP:AMP Phosphotransferase.7 The Functions of Polyphosphates and Polyphosphate-Dependent Enzymes.7.1 Phosphate Reserve.7.1.1 In Prokaryotes.7.1.2 In Eukaryotes.7.2 Energy Source.7.2.1 Polyphosphates in Bioenergetics of Prokaryotes.7.2.2 Polyphosphate in Bioenergetics of Eukaryotes.7.3 Cations Sequestration and Storage.7.3.1 In Prokaryotes.7.3.2 In Eukaryotes.7.4 Participation in Membrane Transport.7.5 Cell Envelope Formation and Function.7.5.1 Polyphosphates in the Cell Envelopes of Prokaryotes.7.5.2 Polyphosphates in the Cell Envelopes of Eukaryotes.7.6 Regulation of Enzyme Activities.7.7 Gene Activity Control, Development and Stress Response.7.7.1 In Prokaryotes.7.7.2 In Lower Eukaryotes.7.8 The Functions of Polyphosphates in Higher Eukaryotes.8 The Peculiarities of Polyphosphate Metabolism in Different Organisms.8.1 Escherichia coli.8.1.1 The Dynamics of Polyphosphates under Culture Growth.8.1.2 The Effects of Pi Limitation and Excess.8.1.3 The Effects of Mutations on Polyphosphate Levels and Polyphosphate-Metabolizing Enzyme Activities.8.1.4 The Effects of Nutrition Deficiency and Environmental Stress.8.2 Pseudomonas aeruginosa.8.3 Acinetobacter.8.4 Aerobacter aerogenes (Klebsiella aerogenes).8.5 Azotobacter.8.6 Cyanobacteria (Blue-Green Algae) and other Photosynthetic Bacteria.8.7 Mycobacteria and Corynebacteria.8.8 Propionibacteria.8.9 Archae.8.10 Yeast.8.10.1 Yeast Cells Possess Different Polyphosphate Fractions.8.10.2 The Dynamics of PolyP Fractions during the Cell Cycle.8.10.3 The Relationship between the Metabolism of Polyphosphates and other Compounds.8.10.4 Polyphosphate Fractions at Growth on a Pi-Sufficient Medium with Glucose.8.10.5 The Effects of Pi Limitation and Excess.8.10.6 The Effects of other Conditions on the Polyphosphate Content in Yeast Cells.8.10.7 The Effects of Inhibitors on the Polyphosphate Content in Yeast Cells.8.10.8 The Effects of Mutations on the Content and Chain Lengths of Polyphosphate in Yeast.8.11 Other Fungi (Mould and Mushrooms).8.12 Algae.8.12.1 Localization and Forms in Cells.8.12.2 The Dynamics of Polyphosphates in the Course of Growth.8.12.3 The Influence of Light and Darkness.8.12.4 The Effects of Pi Limitation and Excess.8.12.5 Changes in Polyphosphate Content under Stress Conditions.8.13 Protozoa.8.14 Higher Plants.8.15 Animals.9 Applied Aspects of Polyphosphate Biochemistry.9.1 Bioremediation of the Environment.9.1.1 Enhanced Biological Phosphate Removal.9.1.2 Removal of Heavy Metals from Waste.9.2 Polyphosphates and Polyphosphate-Metabolizing Enzymes in Assay and Synthesis.9.3 Polyphosphates in Medicine.9.3.1 Antiseptic and Antiviral Agents.9.3.2 Polyphosphate Kinase as a Promising Antimicrobial Target.9.3.3 Polyphosphates as New Biomaterials.9.3.4 Polyphosphates in Bone Therapy and Stomathology.9.4 Polyphosphates in Agriculture.9.5 Polyphosphates in the Food Industry.10 Inorganic Polyphosphates in Chemical and Biological Evolution.10.1 Abiogenic Synthesis of Polyphosphates and Pyrophosphate.10.2 Phosphorus Compounds in Chemical Evolution.10.3 Polyphosphates and Pyrophosphates: Fossil Biochemical Reactions and the Course of Bioenergetic Evolution.10.4 Changes in the Role of Polyphosphates in Organisms at Different Evolutionary Stages.References.Index of Generic Names.Subject Index.

The biochemistry of inorganic polyphosphates

This comprehensive literature review of the phosphorus nutrition and metabolism of eukaryotic microalgae deals sequentially with (1) extracellular P-compounds available for algal utilization and growth; (2) orthophosphate uptake mechanisms, kinetics, and influence from environmental variables; (3) phosphatase-mediated utilization of organic phosphates involving multiple enzymes, induction and cellular location of repressible and irrepressible phosphatases, and their role in growth physiological processes; (4) intracellular phosphate metabolism covering diversity of phosphometabolites. ATP-linked energy regulation, polyphosphate pools and storage roles, phospholipids and phospholipases; (5) steady-state and transient-state models relating phosphate utilization to growth; (6) ecological aspects covering manifestations of phosphorus limitation, interspecific competition for phosphonutrients among microorganisms, and current views on phosphorus cycling and turnover in aquatic ecosystems. Although concentrating on the microalgae, the review often points out sounder conclusions drawn from bacteria and fungi, and includes specific macroalgae in considering certain subtopics where such algae were better investigated and provided a good basis for comparison with the microalgae.

The utilization of inorganic and organic phosphorous compounds as nutrients by eukaryotic microalgae: a multidisciplinary perspective: part 1

SUMMARY

The kinetics of phosphate uptake and growth in Scenedesmus sp. have been studied in continuous culture with particular reference to the shifts in the cellular P compounds as a function of growth rate.



Uptake velocity is a function of both internal and external substrate concentrations and can be described by the kinetics of noncompetitive enzyme inhibition. The concentrations of polyphosphates (alkali-extractable or 7-min) can he substituted as inhibitors in the kinetic equation. The apparent half-saturation constant of uptake. Km, is 0.6 μM. The apparent half-saturation concentration for growth is less than Km, by 1 order of magnitude. Growth is a function of cellular P concentrations, and the polyphosphates (alkali-extractable or 7-min) appear to regulate growth rate directly or indirectly. To understand P limitation, therefore, it is necessary to measure both external P and internal polyphosphate levels. Evidence indicates that alkali-extractable polyphosphates, which can be quantitatively determined by a simple method of measuring surplus P, are involved in cell division process find that a maintenance concentration of functional phosphate exists in the form of poly phosphates. Alkaline phosphatase activity has an inversely linear relationship to growth rate and to the reciprocals of both polyphosphates and surplus P. Changes in lipid P, RNA P, and presumably all other forms except DNA are related to changes in growth rate.

A continuous culture study of phosphate uptake, growth rate and polyphosphate in scenedesmus sp.1

Publisher Summary This chapter discusses the physiological role of inorganic polyphosphates. Inorganic polyphosphates are linear polymers in which orthophosphate residues are linked by energy-rich phospho-anhydride bonds. Inorganic polyphosphates have been found in almost all tested representatives of living cells. They have been detected in eubacteria, fungi, algae, mosses, protozoa, insects, and in various tissues of higher plants and animals. In highly organized organisms, polyphosphates perform several specialized functions, being donors of activated phosphate only for quite specific biochemical or physiological processes. The chapter presents new aspects of inorganic pyrophosphate's metabolic and physiological role. Pyrophosphate is only a byproduct of numerous reactions of pyrophosphorolysis involved in biosynthesis of proteins, nucleic acids, lipids, polysaccharides, and nucleoside coenzymes. The energy-dependent synthesis of pyrophosphate during photosynthetic and oxidative phosphorylation is also discussed. The first evidence for an energy-dependent biosynthesis of pyrophosphate is obtained with animal tissue homogenates. Finally, several modern concepts about the role of high molecular-weight polyphosphates and pyrophosphate in evolution of phosphorus metabolism are presented in the chapter.

Polyphosphate metabolism in micro-organisms

Publisher Summary Microalgae are of very great ecological importance as the major primary producers in large bodies of water. They are also found in benthic habitats, in soil, and in a number of symbioses, particularly corals and lichens. It discusses that microalgae may not seem to be the organisms of choice for investigating the mechanism of the transport of nutrients across the plasmalemma—they are usually photolithotrophs, which poses problems both for the growth of the organisms and for unravelling the energy sources for transport. In addition to the respiratory and fermentative energy sources of heterotrophic cells, there is also the possibility of direct intervention of photoproduced high-energy compounds in powering transport in the light. It has not so far proved easy to separate the plasmalemma from other cell membranes in subcellular preparations, even in the structurally simple cyanobacteria. This is a prerequisite for detailed analysis of membranes and characterization of the transport systems found in them. Their ecological importance and the possibility of testing current chemiosmotic views on the mechanism of membrane transport in organisms, which live at extremes of pH and of osmotic pressure make them very important experimental objects.

Nutrient Transport in Microalgae

Manometric measurements show that oxygen evolution proceeds in synchronised cells of Ankistrodesmus braunii even in an atmosphere of pure nitrogen. In this case the slow oxygen evolution is dependent on the presence of nitrate (Table 1). Light saturation is found at a low light intensity at pH 5.6, at a higher light intensity at pH 8.0 (Fig. 1). The light saturation curves are in good agreement with those of 32P-labelling in Ankistrodesmus under the same conditions (Fig. 2).

[Nitrate-dependent noncyclic photophosphorylation in Ankistrodesmus braunii in the absence of CO2 and O 2].

The effect of CO2 on the 32P-labelling of polyphosphates and acid-soluble organic phosphates is studied in synchronously grown cultures of the green alga Ankistrodesmus braunii, using trichloroacetic acid treatment and acid hydrolysis for the fractionation of the phosphorus compounds. Three per cent CO2 in nitrogen causes an inhibition of the labelling of polyphosphates but a marked increase of 32P in organic phosphates, whereas oxygen (CO2-free air) produces the reverse effect. Polyphosphates and ATP are the fractions most stimulated by O2, while stable organic phosphates show the strongest inhibition. Labelling of nucleic acids is relatively indifferent to both oxygen and CO2. Three per cent CO2 in air causes the same distribution of 32P-labelling as 3 per cent CO2 in N2. 
32P-labelling is strongly dependent on the pH of the medium. In the absence of CO2, polyphosphate labelling is highest in the acidic range, whereas organic phosphates and ATP show optimum labelling and the highest percentage of the total 32P in the alkaline pH range. The effect of CO2 is strongest between pH 5 and 6, that of oxygen between pH 8 and 9. Apparently the pH of the medium exerts a considerable influence upon the phosphate metabolism inside the cells. Increasing concentration of CO2 lead to the same change of 32P-labelling in nitrogen as in air and to saturation at about 1 per cent CO2 under the conditions used. The curves are in good agreement with those of O2-evolution at increasing concentrations of CO2, but they show completely different rates. Young cells respond to CO2 and O2 differently from cells in the photosynthetically most active stage. In young cells both gasses are less effective. The effect of CO2 is explained by a strong increase in noncyclic photophosphorylation which can proceed only slowly in N2. ATP-consumption connected with high rates of CO2-fixation may be the reason for the low rates of 32P-labelling in the polyphosphate fraction when CO2 is present. The influence of external pH on 32P-labelling is partly due to the pH-dependence of phosphate uptake, but the different response of several fractions to the pH of the medium suggests that the pH of the cytoplasm and possibly even the pH of the interior of the chloroplasts is affected by the external pH. The effect of O2 in the absence of CO2 or at low CO2-concentrations is explained by the well-known inhibition of photosynthesis by oxygen. Increasing concentrations of CO2 reverse this inhibition and correspondingly change the distribution of 32P between the phosphate fractions. The change in sensitivity to CO2 and O2 with the cell age is consistent with the change in the rates of maximum photosynthetic CO2-fixation.

[The influence of CO2 and pH on (32)P-labelling of polyphosphates and organic phosphates in Ankistrodesmus braunii in the light].

Durch vergleichende Untersuchung der Transporthemmung und der Kallosebildung in unbehandelten und cyanidvergifteten Zentralleitbundeln der Blattstiele vonPelargonium zonale wurde die Frage einer mechanischen Transporthemmung durch die Kallose gepruft. Cyanid regte in der verwendeten Konzentration die Kallosebildung bei einem Teil der Siebplatten deutlich an; eine Ruckbildung in Leitbundeln, die nach einer Erholungszeit wieder Fluorescein leiteten, wurde nicht beobachtet, doch nahm mit dem Alter der Blattstiele die Kallosemenge auch ohne Vergiftung erheblich zu. Anhand der Ergebnisse wird eine rein mechanische Transporthemmung durch Cyanid infolge von Kallosebildung fur unwahrscheinlich gehalten.

Über die Bildung von Kallose bei einer Hemmung des Transportes in den Siebröhren durch Cyanid

Short-time experiments with 32P-labelled phosphate and chase experiments with equally labelled cells were carried out with synchronized algae under conditions of optimum phosphate uptake. In short-time experiments, in the presence as in the absence of CO2, orthophosphate and organic phosphates are rapidly labelled, but their time curves show saturation behaviour after 10 to 20 min. Labelling of polyphosphates proceeds at a constant rate after a short lag period of about 5 min. In equally labelled algae 32P-labelling correspondingly decreases in orthophosphate and in organic phosphates, but increases by about the same amount in the fraction of acid-insoluble polyphosphates. In the presence of external phosphate and in the light, polyphosphates show no visible decay within the 20 min of the chase experiments. A comparison of the two kinds of experiments suggests that polyphosphates are secondary products of photophosphorylation following only after orthophosphate and organic phosphates, probably after ATP. The rates of photophosphorylation are certainly much higher than the rates of labelling in organic phosphates because of the limiting phosphate uptake. Since the polyphosphates show no decay during the time of the experiments their turnover is low and the rates of polyphosphate labelling after a phosphate starvation period, and after the short lag period, can be regarded as approximate rates of polyphosphate synthesis. These rates are lower than the rates of phosphate uptake. In young cells of the synchronous culture phosphate replenishment after a 5-h starvation requires 2 to 3 h. After replenishment or in a culture undisturbed by phosphate starvation, the rates of polyphosphate accumulation, like the rates of phosphate uptake are much lower. In the presence of CO2 they are constant for several hours, if related to culture volume with constant cell number. Polyphosphate accumulation is proportional to phosphate uptake under these conditions amounting to about one third. In the absence of CO2, the rates decrease after 2 to 4 h of CO2-starvation and, like in short-time experiments a large proportion of the phosphate taken up is used for polyphosphate accumulation. The low rates of long-time experiments may represent a steady state between formation and decay of polyphosphates. Since the cells kept in the absence of CO2 are prevented from growing they actually accumulate more polyphosphates per cell volume, per chlorophyll, and per dry weight than the cells in the presence of CO2. The rates of polyphosphate formation are discussed with respect to their turnover in the light observed by other investigators. They are regarded to be a result of competition for ATP together with the orthophosphate pool of the cells, and of the compartmentation. The rates of polyphosphate formation are rather low compared with the probable rates of ATP formation under various conditions of photophosphorylation. Therefore, the formation of polyphosphates is regarded as a process of secondary order of magnitude in the energy metabolism of algal cells.

Untersuchungen über die Raten der Polyphosphatsynthese durch die Photophosphorylierung bei Ankistrodesmus braunii

Short time incorporation of 32P was carried out with synchronised algae (young cells) depleted of phosphate. For the separation and determination of the acid-insoluble phosphate fractions of the cells an improved fractionation procedure was applied. In order to exclude competition by carbon dioxide all experiments were done in the absence of CO2.

Wolfram R. Ullrich

Papers

[Nitrate-dependent noncyclic photophosphorylation in Ankistrodesmus braunii in the absence of CO2 and O 2].

[The influence of CO2 and pH on (32)P-labelling of polyphosphates and organic phosphates in Ankistrodesmus braunii in the light].

Über die Bildung von Kallose bei einer Hemmung des Transportes in den Siebröhren durch Cyanid

Untersuchungen über die Raten der Polyphosphatsynthese durch die Photophosphorylierung bei Ankistrodesmus braunii

[The effect of oxygen on the (32)P-labelling of polyphosphates and organic phosphates in Ankistrodesmus braunii in the light].