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FEBS Letters Mar 1992In Pseudomonas aeruginosa, the effect of different cations on the acid phosphatase activity was studied in order to acquire more information related to a previously...
In Pseudomonas aeruginosa, the effect of different cations on the acid phosphatase activity was studied in order to acquire more information related to a previously proposed mechanism, involving the coordinated action of this enzyme with phospholipase C. Although the natural substrate of this enzyme is phosphorylcholine, in order to avoid the possible interaction of its positive charge and those of the different cations with the enzyme molecule, the artificial substrate p-nitrophenylphosphate was utilized. Kinetic studies of the activation of acid phosphatase (phosphorylcholine phosphatase) mediated by divalent cations Mg2+, Zn2+ and Cu2+ revealed that all these ions bind to the enzyme in a compulsory order (ordered bireactant system). The Km values obtained for p-NPP in the presence of Mg2+, Zn2+ and Cu2+ were 1.4 mM, 1.0 mM and 3.5 mM, respectively. The KA values for the same ions were 1.25 mM, 0.05 mM and 0.03 mM, respectively. The Vmax obtained in the presence of Cu2+ was about twofold higher than that obtained in the presence of Mg2+ or Zn2+. The inhibition observed with Al3+ seems to be a multi-site inhibition. The K'app and n values, from the Hill plot, were about 0.25 mM and 4.0 mM, respectively, which were independent of the metal ion utilized as activator. It is proposed that the acid phosphatase may exert its action under physiological conditions, depending on the availability of either one of these metal ions.
Topics: Acid Phosphatase; Aluminum; Cations, Divalent; Copper; Enzyme Activation; Kinetics; Magnesium; Pseudomonas aeruginosa; Zinc
PubMed: 1544481
DOI: 10.1016/0014-5793(92)80108-s -
Plant, Cell & Environment Jun 2019Arbuscules are the central structures of the symbiotic association between terrestrial plants and arbuscular mycorrhizal (AM) fungi. However, arbuscules are also...
Arbuscules are the central structures of the symbiotic association between terrestrial plants and arbuscular mycorrhizal (AM) fungi. However, arbuscules are also ephemeral structures, and following development, these structures are soon digested and ultimately disappear. Currently, little is known regarding the mechanism underlying the digestion of senescent arbuscules. Here, biochemical and functional analyses were integrated to test the hypothesis that a purple acid phosphatase, GmPAP33, controls the hydrolysis of phospholipids during arbuscule degeneration. The expression of GmPAP33 was enhanced by AM fungal inoculation independent of the P conditions in soybean roots. Promoter-β-glucuronidase (GUS) reporter assays revealed that the expression of GmPAP33 was mainly localized to arbuscule-containing cells during symbiosis. The recombinant GmPAP33 exhibited high hydrolytic activity towards phospholipids, phosphatidylcholine, and phosphatidic acid. Furthermore, soybean plants overexpressing GmPAP33 exhibited increased percentages of large arbuscules and improved yield and P content compared with wild-type plants when inoculated with AM fungi. Mycorrhizal RNAi plants had high phospholipid levels and a large percentage of small arbuscules. These results in combination with the subcellular localization of GmPAP33 at the plasma membrane indicate that GmPAP33 participates in arbuscule degeneration during AM symbiosis via involvement in phospholipid hydrolysis.
Topics: Acid Phosphatase; Cell Membrane; Gene Expression Regulation, Plant; Glucuronidase; Mycorrhizae; Phosphates; Phospholipids; Plant Proteins; Plant Roots; Plants, Genetically Modified; RNA Interference; Recombinant Proteins; Glycine max; Symbiosis
PubMed: 30730567
DOI: 10.1111/pce.13530 -
The Journal of Biological Chemistry Nov 1980Acid phosphatase-1 (orthophosphoric monoester phosphohydrolase, acid optimum, EC 3.1.3.2), the major phosphatase in adult Drosophila melanogaster, has been purified to...
Acid phosphatase-1 (orthophosphoric monoester phosphohydrolase, acid optimum, EC 3.1.3.2), the major phosphatase in adult Drosophila melanogaster, has been purified to apparent homogeneity. The final product is a glycoprotein homodimer with a subunit molecular weight of about 50,000, as measured by its electrophoretic mobility in denaturing conditions on polyacrylamide gels containing sodium dodecyl sulfate. It has a turnover number of 1720 1-naphthyl phosphate molecules hydrolyzed/s by each acid phosphatase-1 molecule at 37 degrees C, pH 5.0. An average fly contains about 5 ng of enzyme. Pure acid phosphatase-1 displays heterogeneity in isoelectric focusing, with a major band at pH 5.3. The enzyme hydrolyzes a wide variety of phosphate monoesters, including AMP, glucose 6-phosphate, ATP, choline phosphate, or phosphoproteins. The maximum reaction rates are different for all substrates, and some substrates appear to inhibit the reaction at high substrate concentrations. The Michaelis constants for 1-naphthyl phosphate and p-nitrophenyl phosphate are 79 microM and 68 microM, respectively, at pH 5.0 and 37 degrees C. The optimum pH level for 1-naphthyl phosphate is 4.5. Acid phosphatase-1 is inhibited by L(+)-tartrate (but not D(-)-tartrate), phosphate, and fluoride. The reaction rate increases 2.1-fold for every 10 degrees C rise in temperature. Above 48 degrees C, the rate of thermal denaturation is greater than the rate of the enzyme reaction.
Topics: Acid Phosphatase; Animals; Drosophila melanogaster; Isoenzymes; Kinetics; Macromolecular Substances; Molecular Weight
PubMed: 6776114
DOI: No ID Found -
Biochemical and Biophysical Research... Apr 1999Bacterial strains were isolated from the pig colon to screen for phytase and acid phosphatase activities. Among 93 colonies, Colony 88 had the highest activities for...
Bacterial strains were isolated from the pig colon to screen for phytase and acid phosphatase activities. Among 93 colonies, Colony 88 had the highest activities for both enzymes and was identified as an Escherichia coli strain. Using primers derived from the E. coli pH 2.5 acid phosphatase appA sequence (Dassa et al. (1990), J. Bacteriol. 172, 5497-5500), we cloned a 1482 bp DNA fragment from the isolate. In spite of 95% homology between the sequenced gene and the appA, 7 amino acids were different in their deduced polypeptides. To characterize the properties and functions of the encoded protein, we expressed the coding region of the isolated DNA fragment and appA in Pichia pastoris, respectively, as r-appA2 and r-appA. The recombinant protein r-appA2, like r-appA and the r-phyA phytase expressed in Aspergillus niger, was able to hydrolyze phosphorus from sodium phytate and p-nitrophenyl phosphate. However, there were distinct differences in their pH profiles, Km and Vmax for the substrates, specific activities of the purified enzymes, and abilities to release phytate phosphorus in soybean meal. In conclusion, the DNA fragment isolated from E. coli in pig colon seems to encode for a new acid phosphatase/phytase and is designated as E. coli appA2.
Topics: 6-Phytase; Acid Phosphatase; Amino Acid Sequence; Animals; Base Sequence; Cloning, Molecular; Colon; Escherichia coli; Escherichia coli Proteins; Fungi; Genes, Bacterial; Glycosylation; Hydrogen-Ion Concentration; Kinetics; Molecular Sequence Data; Molecular Weight; Multienzyme Complexes; Nitrophenols; Organophosphorus Compounds; Phosphorus; Phytic Acid; Recombinant Proteins; Sequence Homology, Amino Acid; Swine; Temperature
PubMed: 10092520
DOI: 10.1006/bbrc.1999.0361 -
Basic & Clinical Pharmacology &... Feb 2011Snake venom components, acting in concert in the prey, cause their immobilization and initiate digestion. To achieve this, several hydrolytic enzymes of snake venom have... (Review)
Review
Snake venom components, acting in concert in the prey, cause their immobilization and initiate digestion. To achieve this, several hydrolytic enzymes of snake venom have evolved to interfere in various physiological processes, which are well defined. However, hydrolytic enzymes such as phosphatases (acid and alkaline phosphomonoesterases) are less studied and their pharmacological role in venoms is not clearly defined. Also, they show overlapping substrate specificities and have other common biochemical properties causing uncertainty about their identity in venoms. The near-ubiquitous distribution of these enzymes in venoms, suggests a significant role for these enzymes in envenomation. It appears that these enzymes may play a central role in liberating purines (mainly adenosine) - a multitoxin and through the action of purines help in prey immobilization. However, apart from this, these enzymes could also possess other pharmacological activities as venom enzymes have been evolved to interfere in diverse physiological processes. This has not been verified by pharmacological studies using purified enzymes. Further research is needed to biologically characterize these enzymes in snake venoms, such that their role in venom is clearly established.
Topics: Acid Phosphatase; Adenosine; Alkaline Phosphatase; Animals; Snake Venoms; Snakes
PubMed: 21156030
DOI: 10.1111/j.1742-7843.2010.00630.x -
Journal of Bacteriology May 1990One of the signals that has been reported to be important in stimulating fruiting body formation of Myxococcus xanthus is starvation for phosphate. We therefore chose to...
One of the signals that has been reported to be important in stimulating fruiting body formation of Myxococcus xanthus is starvation for phosphate. We therefore chose to study phosphatase activity during M. xanthus development. Many phosphatases can cleave the substrate p-nitrophenol phosphate. Using this substrate in buffers at various pHs, we obtained a profile of phosphatase activities during development and germination of M. xanthus. These experiments indicated that there are five patterns of phosphatase activity in M. xanthus: two vegetative and three developmental. The two uniquely vegetative activities have pH optima at 7.2 and 8.5. Both require magnesium and both are inhibited by the reducing agent dithiothreitol. The developmental (spores) patterns of activity have pH optima of 5.2, 7.2, and 8.5. All three activities are Mg independent. Only the alkaline phosphatase activity is inhibited by dithiothreitol. The acid phosphatase activity is induced very early in development, within the first 2 to 4 h. Both the neutral and alkaline phosphatase Mg-independent activities are induced much later, about the time that myxospores become evident (24 to 30 h). The three activities are greatly diminished upon germination; however, the kinetics of loss differ for all three. The acid phosphatase activity declines very rapidly, the neutral activity begins to decline only after spores begin to convert to rods, and the alkaline phosphatase activity remains high until the time the cells begin to divide. All three developmental activities were measured in the developmental signalling mutants carrying asg, csg, and dsg. The pattern of expression obtained in the mutants was consistent with that of other developmentally regulated genes which exhibit similar patterns of expression during development. The ease with which phosphatases can be assayed should make the activities described in this report useful biochemical markers of stages of both fruiting body formation and germination.
Topics: Acid Phosphatase; Alkaline Phosphatase; Enzyme Induction; Genotype; Hydrogen-Ion Concentration; Kinetics; Myxococcales; Phosphoric Monoester Hydrolases
PubMed: 2158968
DOI: 10.1128/jb.172.5.2294-2302.1990 -
The Journal of Experimental Medicine Nov 1968The acid phosphatase activity of normal alveolar and BCG-induced alveolar macrophages has been examined. Five electrophoretically distinct forms of acid phosphatase have...
The acid phosphatase activity of normal alveolar and BCG-induced alveolar macrophages has been examined. Five electrophoretically distinct forms of acid phosphatase have been identified in both normal and BCG-induced macrophages. The acid phosphatases can be divided into two major categories. One category, containing four distinct forms, is readily solubilized after repeated freezing and thawing or mechanical disruption The second category, containing one form, is firmly bound to the lysosomal membrane and can be solubilized by treatment of the lysosomal fraction with Triton X-100. The Triton-extractable acid phosphatase and the predominant aqueous soluble acid phosphatase have been shown to differ in the degree of membrane binding, in solubility, in net charge, and in molecular weight. The two pre-dominant phosphatases possess identical pH optimum and do not differ in response to enzyme inhibitors. BCG stimulation has been shown to result in a nearly twofold increase in acid phosphatase activity. A nearly proportionate increase in the major acid phosphatase forms has been observed.
Topics: Acid Phosphatase; Animals; Antigens; BCG Vaccine; Chromatography, Gel; Chromatography, Ion Exchange; Electrophoresis; Hydrogen-Ion Concentration; Isoenzymes; Lysosomes; Macrophages; Molecular Weight; Mycobacterium bovis; Rabbits; Tartrates
PubMed: 4878908
DOI: 10.1084/jem.128.5.1031 -
Journal of Biochemistry Jan 1984An acid phosphatase species which is activated by Fe2+ was purified 3,700-fold from rat spleen by chromatography on columns containing Blue-Sepharose, concanavalin...
An acid phosphatase species which is activated by Fe2+ was purified 3,700-fold from rat spleen by chromatography on columns containing Blue-Sepharose, concanavalin A-Sepharose, Sephadex G-100, and CM-Sephadex. The enzyme hydrolyzed aryl phosphates, nucleoside di- and triphosphates, phosphoproteins, and thiamine pyrophosphate with Km values of 10(-4) to 10(-3) M at an optimal pH of 5.0-5.8. Co-purification of the acid phosphatase and acid phosphoprotein phosphatase indicated that they were identical. The purified enzyme was glycoprotein in nature, showing four heterogeneous forms on acid polyacrylamide gel electrophoresis (pI values, 7.8, 8.0, 8.3, and 8.5), but it gave a molecular weight of 33,000 on sodium dodecyl sulfate-gel electrophoresis and gel permeation chromatography. The enzyme had a purple color (lambda max 545 nm) and contained 2 iron atoms per enzyme molecule. Among reductants, ascorbic acid and Fe2+ were the best activators, although their combined effect was not additive. Fe2+ and ascorbic acid both changed the purple enzyme into the same active form (lambda max 515 nm), giving almost the same kinetic constants for substrates and for inhibitors such as molybdate, phosphate and fluoride. However, low concentrations of Fe2+, from 0.01 mM to 1.0 mM, immediately and reversibly activated the enzyme, whereas high concentrations of ascorbic acid over 1 mM were required for maximal activation, which was slow and irreversible.
Topics: Acid Phosphatase; Animals; Chemical Phenomena; Chemistry; Electrophoresis, Polyacrylamide Gel; Enzyme Activation; Iron; Male; Rats; Spleen; Substrate Specificity
PubMed: 6706919
DOI: 10.1093/oxfordjournals.jbchem.a134603 -
The Journal of Biological Chemistry Sep 1994At the active site of rat prostatic acid phosphatase (rPAP), residue Asp258 is a suitable candidate to act as an acid/base catalyst during phosphoester hydrolysis. It... (Comparative Study)
Comparative Study
At the active site of rat prostatic acid phosphatase (rPAP), residue Asp258 is a suitable candidate to act as an acid/base catalyst during phosphoester hydrolysis. It was changed to Asn, Ser, and Ala by site-directed mutagenesis. All these mutants were inactive, indicating that Asp258 may act as a proton donor in catalysis. Tyr123 and Arg127 residues, located at the entrance of the active site surface in rPAP, are likely to be responsible for the substrate specificity of the enzyme. The corresponding residues in lysosomal acid phosphatase (LAP) are Lys and Gly. In order to clarify the roles of the Tyr123 and Arg127 residues, lysosomal type rPAP mutants (Y123K, R127G and Y123K,R127G) were generated. Sensitivity of Y123K,R127G to tartrate inhibition was similar to that observed in the case of LAP, indicating that these residues might be responsible for differences in substrate specificity between the enzymes of prostatic and lysosomal origin. However, unlike human LAP, the lysosomal type mutants hydrolyzed the suggested PAP-specific substrates, phosphocreatine and phosphocholine, showing that Tyr123 and Arg127 are not the only residues contributing to the substrate specificity of rPAP. The residues Trp106 and His112 appeared to be important in the dimerization of rPAP. Oligomerization mutants (W106E, H112D and W106E,H112D) existed in a monomeric form without catalytic activity or a tartrate binding ability.
Topics: Acid Phosphatase; Amino Acid Sequence; Animals; Aspartic Acid; Baculoviridae; Base Sequence; DNA Primers; Electrophoresis, Polyacrylamide Gel; Hydrogen-Ion Concentration; Immunoblotting; Kinetics; Macromolecular Substances; Male; Molecular Sequence Data; Moths; Mutagenesis, Site-Directed; Point Mutation; Polymerase Chain Reaction; Prostate; Rats; Recombinant Proteins; Substrate Specificity; Transfection
PubMed: 8077215
DOI: No ID Found -
Progress in Biophysics and Molecular... 2000
Comparative Study Review
Topics: Acid Phosphatase; Alkaline Phosphatase; Amino Acid Sequence; Binding Sites; Evolution, Molecular; Geobacillus stearothermophilus; Models, Molecular; Molecular Sequence Data; Phosphofructokinase-2; Phosphoglycerate Mutase; Phosphoric Monoester Hydrolases; Protein Structure, Secondary; Saccharomyces cerevisiae; Sequence Homology, Amino Acid
PubMed: 10958932
DOI: 10.1016/s0079-6107(00)00007-9