Activation of folylpolyglutamate synthetase by pteroic acid

1985 ◽  
Vol 63 (7) ◽  
pp. 777-779 ◽  
Author(s):  
P. J. Vickers ◽  
R. Di Cecco ◽  
Z. B. Pristupa ◽  
K. G. Scrimgeour
Keyword(s):  
Conformational Change ◽  
Ph Optimum ◽  
Beef Liver ◽  
Folylpolyglutamate Synthetase ◽  
Ph Values ◽  
Neutral Ph ◽  
Pteroic Acid ◽  
Stimulation Of

The glutamylation of methotrexate catalyzed by beef liver folylpolyglutamate synthetase (FPGS) is activated by addition of pteroic acid. Pteroic acid causes greater stimulation of FPGS, including glutamylation of tetrahydrofolate, at neutral pH values (i.e., below the pH optimum of 8.4). We have attributed this activation to a conformational change of FPGS induced by pteroic acid.

Biosynthesis of polyglutamates of folates

1986 ◽  
Vol 64 (7) ◽  
pp. 667-674 ◽  
Author(s):  
K. G. Scrimgeour
Keyword(s):  
Current Knowledge ◽  
Competitive Inhibitor ◽  
High Dose ◽  
Beef Liver ◽  
Folylpolyglutamate Synthetase ◽  
Low Concentrations ◽  
Alternate Substrate ◽  
Pteroic Acid ◽  
Rescue Agent ◽  
Addition Pathway

Folylpolyglutamate synthetase (FPGS) catalyzes the synthesis of the poly-γ-glutamate forms of tetrahydrofolate and its coenzyme adducts, as well as of the folate-analogue drugs. This paper reviews current knowledge of the preparations of FPGS from mammalian sources (rat, hog, mouse, and beef liver). Kinetic constants for the substrates and activators of FPGS are compared. Tetrahydrofolate and 5-formyltetrahydrofolate are excellent substrates for the enzyme. The Km values for the antifolates and their 7-hydroxy metabolites are much higher than those for the tetrahydrofolates. Aminopterin has higher activity with FPGS than does methotrexate, which partially explains its greater toxicity. 5-Formyltetrahydrofolate, which is used as a rescue agent in high-dose methotrexate-rescue chemotherapy, is a better alternate substrate of FPGS than is methotrexate and therefore is a potent competitive inhibitor of the glutamylation of methotrexate. Thus, low concentrations of the rescue agent prevent formation of cytotoxic polyglutamates of methotrexate. The pathway of the reaction is the addition of a glutamate residue to the terminal γ-carboxyl of the pteridine substrate. That longer folylpolyglutamates are poorer substrates possibly is a result of this addition pathway. Pteroic acid activates FPGS by lowering the Km value of the pteridine substrate. It also greatly increases the activity of the synthetase at physiological pH values.

scholarly journals Identification and characterization of a phospholipase C activity in resident mouse peritoneal macrophages. Inhibition of the enzyme by phenothiazines

1981 ◽  
Vol 197 (2) ◽  
pp. 523-526 ◽  
Author(s):  
Paul D. Wightman ◽  
Mary Ellen Dahlgren ◽  
James C. Hall ◽  
Philip Davies ◽  
Robert J. Bonney
Keyword(s):  
Phospholipase C ◽  
High Activity ◽  
Peritoneal Macrophages ◽  
Ph Optimum ◽  
Reaction Velocity ◽  
Neutral Ph ◽  
Maximum Reaction ◽  
Mouse Peritoneal Macrophages ◽  
Identification And Characterization

Resident mouse peritoneal macrophages contain a phospholipase C of high activity that is specific for phosphatidylinositol. The activity has a neutral pH optimum, is Ca2+-dependent and has a maximum reaction velocity of 525nmol/h per mg of protein. Certain phenothiazines are potent inhibitors of this activity.

scholarly journals The Listeria monocytogenes hemolysin has an acidic pH optimum to compartmentalize activity and prevent damage to infected host cells

2002 ◽  
Vol 156 (6) ◽  
pp. 1029-1038 ◽  
Author(s):  
Ian J. Glomski ◽  
Margaret M. Gedde ◽  
Albert W. Tsang ◽  
Joel A. Swanson ◽  
Daniel A. Portnoy
Keyword(s):  
Listeria Monocytogenes ◽  
Bacterial Pathogen ◽  
Cytolytic Activity ◽  
Host Cells ◽  
Adaptive Mutation ◽  
Acidic Ph ◽  
Listeriolysin O ◽  
Lysosomal Hydrolases ◽  
Ph Optimum ◽  
Neutral Ph

Listeria monocytogenes is a facultative intracellular bacterial pathogen that escapes from a phagosome and grows in the host cell cytosol. The pore-forming cholesterol-dependent cytolysin, listeriolysin O (LLO), mediates bacterial escape from vesicles and is ∼10-fold more active at an acidic than neutral pH. By swapping dissimilar residues from a pH-insensitive orthologue, perfringolysin O (PFO), we identified leucine 461 as unique to pathogenic Listeria and responsible for the acidic pH optimum of LLO. Conversion of leucine 461 to the threonine present in PFO increased the hemolytic activity of LLO almost 10-fold at a neutral pH. L. monocytogenes synthesizing LLO L461T, expressed from its endogenous site on the bacterial chromosome, resulted in a 100-fold virulence defect in the mouse listeriosis model. These bacteria escaped from acidic phagosomes and initially grew normally in cells and spread cell to cell, but prematurely permeabilized the host membrane and killed the cell. These data show that the acidic pH optimum of LLO results from an adaptive mutation that acts to limit cytolytic activity to acidic vesicles and prevent damage in the host cytosol, a strategy also used by host cells to compartmentalize lysosomal hydrolases.

scholarly journals Immunocytochemical localization and biochemical characterization of a novel plasma membrane-associated, neutral pH optimum α-l-fucosidase from rat testis and epididymal spermatozoa

1996 ◽  
Vol 318 (3) ◽  
pp. 821-831 ◽  
Author(s):  
Manuel AVILÉS ◽  
Irene ABASCAL ◽  
José Angel MARTÍNEZ-MENÁRGUEZ ◽  
María Teresa CASTELLS ◽  
Sheri R. SKALABAN ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Biochemical Characterization ◽  
Gradient Centrifugation ◽  
Light And Electron Microscopy ◽  
Enzymic Activity ◽  
Epididymal Sperm ◽  
Sucrose Density Gradient Centrifugation ◽  
Ph Optimum ◽  
Neutral Ph

1. Immunocytochemical and biochemical techniques have been used to localize and characterize a novel plasma membrane-associated, neutral-pH-optimum α-l-fucosidase from rat spermatozoa. Light and electron microscopy specifically localized the fucosidase on the plasma membrane of the convex region of the principal segment of testicular and cauda epididymal sperm heads. Immunoreactivity for α-l-fucosidase was also detected in the Golgi apparatus of spermatocytes and spermatids but no immunoreactivity was observed in the acrosome. 2. Fractionation of epididymal sperm homogenates indicated that over 90% of the α-l-fucosidase activity was associated with the 48000 g pellet. This pellet-associated activity could be solubilized with 0.5 M NaCl but not with 0.5% Triton X-100, suggesting that fucosidase is peripherally associated with membranes. Sucrose-density-gradient centrifugation of sperm homogenates indicated that fucosidase was enriched in the plasma membrane-enriched fraction. Analysis of α-l-fucosidase on intact epididymal sperm indicated that the enzyme was active, displayed linear kinetics and had a pH–activity curve (with an optimum near 7) which was comparable to that of fucosidase from epididymal sperm extracts. These results further suggest that fucosidase is associated with plasma membranes, and that its active site is accessible to fucoconjugates. Evidence that most of the fucosidase is associated with the exterior of the plasma membrane came from studies in which intact sperm had fucosidase activity comparable to that of sperm sonicates, and from studies in which approx. 90% of the fucosidase activity on intact sperm could be released from the sperm by gentle shaking with 0.5 M NaCl. Isoelectric focusing indicated that the NaCl-solubilized epididymal sperm fucosidase appears to have one major and one minor isoform with pIs near 7.2 and 5.2, respectively. SDS/PAGE and Western blotting indicated that the NaCl-solubilized extract of epididymal sperm contains two protein bands of 54 and 50 kDa which were highly immunoreactive with the IgG fraction of anti-fucosidase antibodies. Although the function of the novel sperm fucosidase is not known, its specific localization to the plasma membrane of the region of the rat sperm head involved in sperm–egg binding and its high enzymic activity at neutral pH on intact sperm suggest that this enzyme may have a role in sperm–egg interactions.

scholarly journals Comparison between Pseudomonas aeruginosa siderophores and desferrioxamine for iron acquisition from ferritin

2010 ◽  
Vol 4 (4) ◽  
pp. 631-635 ◽  
Author(s):  
Somporn Srifuengfung ◽  
Susan Assanasen ◽  
Malulee Tuntawiroon ◽  
Sumonrat Meejanpetch
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Iron Acquisition ◽  
Iron Chelator ◽  
Dialysis Membrane ◽  
In Vitro Experiment ◽  
Iron Mobilization ◽  
Ph Values ◽  
Minimum Medium ◽  
Stimulation Of

Abstract Background: Siderophore is an iron chelator produced by microorganism. Pseudomonas aeruginosa produces two siderophores (pyoverdin and pyochelin). Desferrioxamine is a siderophore used in thalassemia patients to treat an iron overload of vital organs. Objective: Compare the ability of pyoverdin, pyochelin, and desferrioxamine for iron mobilization from ferritin. Materials and Methods: In vitro experiment, the ability of P. aeruginosa siderophores and desferrioxamine for iron mobilization from ferritin was compared by using a dialysis membrane assay at pH values of 7.4 and 6.0. Stimulation of P. aeruginosa PAO1 growth by all siderophores was studied in glucose minimum medium. Results: All three compounds were capable of iron mobilization at both pHs. At pH 6.0, the most effectiveness compound was desferrioxamine (31.6%), followed by pyoverdin (21.5%) and pyochelin (13.7%) compared on weight basis, each at 10 μg/mL. At equimolar concentration, their activities were desferrioxamine (38.5±1.2%), followed by pyoverdin (32.0±4.8%) and pyochelin (26.7±1.9%), respectively. Conclusion: The most effective compound in iron mobilization from ferritin was desferrioxamine, followed by pyoverdin and pyochelin respectively.

Soluble neutral maltase–glucoamylase from the small intestine: separation and characterization of components with differing affinity for concanavalin A

1982 ◽  
Vol 60 (11) ◽  
pp. 1007-1013 ◽  
Author(s):  
G. Forstner ◽  
A. Salvatore ◽  
L. Lee ◽  
J. Forstner
Keyword(s):  
Concanavalin A ◽  
Gradient Elution ◽  
Soluble Form ◽  
Rat Intestine ◽  
Ph Optimum ◽  
Molecular Weights ◽  
Neutral Ph ◽  
Membrane Bound ◽  
Sepharose 4B

Intestinal maltase with a neutral pH optimum exists in both a brush border membrane-bound form and a soluble form in suckling rat intestine. Previous experiments in our laboratory have shown that the soluble enzyme contains a component which binds much more tightly to concanavalin A (ConA) than solubilized forms of the membrane enzyme. We studied the origin of this component by subjecting neutral, soluble maltase activity to chromatography on Sepharose 4B at age 13, 18 (preweaning), and 25 (postweaning) days. At 13 days, two maltase peaks were obtained with approximate molecular weights of 400 000 (peak I) and 150 000 (peak II). Peak II was less prominent at 18 days and was absent at 25 days. At 13 days, the majority of peak I consisted of material which was bound between 0.025 and 0.05 M α-methyl mannoside on gradient elution chromatography of ConA-Sepharose. Peak II contained material which eluted between 0.075 and 0.3 M α-methyl mannoside. At 25 days, all of the soluble maltase eluted between 0.025 and 0.04 M α-methyl mannoside. Peak I and peak II maltases had similar pH optima and Km's for maltase. Peak II maltase had a fourfold greater activity toward glycogen than peak I maltase with approximately the same activity for palatinose, turanose, and trehalose. Both maltases were precipitated by an antibody raised against adult membrane-bound maltase. Soluble maltase with neutral pH activity in the suckling rat intestine, therefore, consists of two immunologically related isozymes which differ in their molecular weight, their binding by ConA, and their specificity for glycogen. The small isozyme disappears at or about the time of weaning.

scholarly journals Separation and properties of two arylamidases from rat cardiac-muscle extracts

1977 ◽  
Vol 163 (3) ◽  
pp. 565-570 ◽  
Author(s):  
A F Bury ◽  
T Coolbear ◽  
C R Savery
Keyword(s):  
Metal Ions ◽  
Cardiac Muscle ◽  
Rat Heart ◽  
Bivalent Metal ◽  
Ph Optimum ◽  
Minor Peak ◽  
Bivalent Metal Ions ◽  
Stimulation Of

Two main arylamidase activities were separated from a particle-free supernatant of rat heart by chromatography on DEAE-Sephadex. Although both enzymes hydrolysed L-leucine 4-nitroanilide, only peak-II enzyme hydrolysed L-lysine 4-nitroanilide. A third minor peak (Ia) contained an enzyme that was active mainly on the L-lysine 4-nitroanilide. The mol.wts. of the enzymes in peaks I and II were approx. 257000 and 105000 respectively. The pH optimum was approx. pH7.0 for peak-I enzyme and 7.0-8.0 for peak-II enzyme. Both enzymes were inhibited by addition of puromycin, p-hydroxymercuribenzoate, o-phenanthroline and bivalent metal ions. Addition of dithiothreitol resulted in stimulation of both activities. Dialysis against o-phenanthroline resulted in inhibition of peak-I and -II enzymes, but after dialysis against EDTA only peak-II enzyme was inhibited.

scholarly journals Inactivation of tyrosine aminotransferase in neutral homogenates and rat liver slices

1972 ◽  
Vol 129 (5) ◽  
pp. 1131-1138 ◽  
Author(s):  
F. Auricchio ◽  
L. Mollica ◽  
A. Liguori
Keyword(s):  
Amino Acid ◽  
Rat Liver ◽  
Acid Concentration ◽  
Tyrosine Aminotransferase ◽  
Amino Acid Concentration ◽  
Acidic Ph ◽  
Ph Values ◽  
Neutral Ph ◽  
Liver Slices

Inactivation of tyrosine aminotransferase induced in vivo by triamcinolone was studied in a homogenate incubated at neutral pH values. The integrity and the presence of subcellular particles together with a compartment of acidic pH are necessary for inactivation of tyrosine aminotransferase. It is suggested that tyrosine aminotransferase is inactivated inside lysosomes. The system responsible for inactivation of tyrosine aminotransferase was partially purified and identified with lysosomal cathepsins B and B1. Inactivation of tyrosine aminotransferase in liver slices is controlled by the amino acid concentration and strongly stimulated by cysteine. 3,3′,5-Tri-iodo-l-thyronine reversibly and strongly decreases the rate of inactivation of tyrosine aminotransferase. The effect is not due to an increased rate of tyrosine aminotransferase synthesis.

Thermostable arginase from Sulfobacillus acidophilus with neutral pH optimum applied for high-efficiency l-ornithine production

2020 ◽  
Vol 104 (15) ◽  
pp. 6635-6646
Author(s):  
Kai Huang ◽  
Shurong Zhang ◽  
Xiao Guan ◽  
Jing Liu ◽  
Sen Li ◽  
...  
Keyword(s):  
High Efficiency ◽  
Ph Optimum ◽  
Neutral Ph
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