scholarly journals Alkaline phosphatase from pig kidney. Microheterogeneity and the role of neuraminic acid

1974 ◽  
Vol 141 (1) ◽  
pp. 293-298 ◽  
Author(s):  
Kunio Hiwada ◽  
Ernst D. Wachsmuth
Keyword(s):  
Alkaline Phosphatase ◽  
Brush Border Membrane ◽  
Deae Cellulose ◽  
Multiple Forms ◽  
Alkaline Phosphatases ◽  
Acetylneuraminic Acid ◽  
Pig Kidney ◽  
Ph Dependency ◽  
Polypeptide Chains

Several alkaline phosphatases (EC 3.1.3.1) could be obtained from pig kidney brush-border membrane on extraction with butan-1-ol. Three of the multiple forms were separated by DEAE-cellulose chromatography and further purified. They form a regular series with different degrees of glycosylation (mainly owing to N-acetylneuraminic acid), of charge, of molecular weight, of stability to temperature, to pH and to urea, of minimal requirement for Mg2+ and of extractability by butan-1-ol. In contrast, the detectable antigenic sites, the inhibition by amino acids and the pH-dependency of Km and Vmax. were identical for these multiple forms. On treatment with neuraminidase, the multiple forms became identical in all their properties. It was therefore concluded that the microheterogeneity of alkaline phosphatase is due to different degrees of glycosylation at polypeptide chains which appear to be otherwise identical.

scholarly journals Role of neuraminic acid in the heterogeneity of alkaline phosphatase in sheep brain

1968 ◽  
Vol 107 (2) ◽  
pp. 185-190 ◽  
Author(s):  
S. Saraswathi ◽  
B. K. Bachhawat
Keyword(s):  
Alkaline Phosphatase ◽  
Deae Cellulose ◽  
Acetylneuraminic Acid ◽  
Deae Cellulose Column ◽  
Elution Pattern ◽  
Sheep Brain ◽  
Substrate Affinities ◽  
Enzyme I ◽  
Cellulose Column

1. Two alkaline phosphatase fractions from sheep brain obtained by DEAE-cellulose column chromatography were shown to be associated with different concentrations of NANA (N-acetylneuraminic acid). Enzyme II contains nearly three times as much NANA as does enzyme I. 2. Partial removal of NANA by neuraminidase digestion from these alkaline phosphatase fractions has different effects on their chromatographic properties. Though the enzymic release of NANA has no effect on the elution pattern of enzyme I from a DEAE-cellulose column, such a treatment shifts the elution pattern of enzyme II towards that of enzyme I. 3. However, this change in the elution pattern of enzyme II as a result of the removal of NANA does not produce any change in the kinetics of this fraction, and the differences between enzyme I and enzyme II with respect to their substrate affinities and Ki for phosphate inhibition are maintained even after the removal of NANA. 4. Results indicate that NANA is not the only factor responsible for the heterogeneity of alkaline phosphatase in sheep brain and enzyme I is not the result of the removal of NANA from enzyme II.

Role of Alkaline Phosphatase in Phosphate Uptake into Brush Border Membrane Vesicles from Human Intestinal Mucosa

1985 ◽  
Vol 97 (5) ◽  
pp. 1461-1466 ◽  
Author(s):  
Kazuyuki HIRANO ◽  
Yuichi IIIZUMI ◽  
Yukio MORI ◽  
Kazumi TOYOSHI ◽  
Mamoru SUGIURA ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Intestinal Mucosa ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Phosphate Uptake ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles

scholarly journals Alkaline phosphatase from pig kidney. Method of purification and molecular properties

1974 ◽  
Vol 141 (1) ◽  
pp. 273-282 ◽  
Author(s):  
Ernst D. Wachsmuth ◽  
Kunio Hiwada
Keyword(s):  
Alkaline Phosphatase ◽  
Brush Border ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Molecular Properties ◽  
Purification Procedure ◽  
Brush Border Membranes ◽  
Pig Kidney ◽  
Critical Ph

Alkaline phosphatase (EC 3.1.3.1) from pig kidney brush-border membranes was solubilized from membrane precipitates by butan-1-ol at a critical pH of 7.0. The 12000-fold purification procedure included (NH4)2SO4 precipitation, DEAE-and TEAE-cellulose chromatography, Sephadex G-200 gel filtration and neuraminidase digestion followed by DEAE-cellulose chromatography. The purified protein contained 20% (w/w) carbohydrate and had mol.wt. 150000–156000 as estimated by Sephadex filtration and ultracentrifuge analysis. It was a tetrameric glycoprotein consisting of identical subunits, and it had a molecular activity at 25°C of 2600s-1 per tetramer. Its concentration in kidney was estimated to be 8.5–8.8mg/kg.

Crystal structure of rat intestinal alkaline phosphatase – Role of crown domain in mammalian alkaline phosphatases

2013 ◽  
Vol 184 (2) ◽  
pp. 182-192 ◽  
Author(s):  
Kaushik Ghosh ◽  
Debarati Mazumder Tagore ◽  
Rushith Anumula ◽  
Basanth Lakshmaiah ◽  
P.P.B.S. Kumar ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Alkaline Phosphatase ◽  
Intestinal Alkaline Phosphatase ◽  
Alkaline Phosphatases

Integumental phosphatase isoenzymes from white puparia of Ceratitis capitata: isolation and characterization

1991 ◽  
Vol 69 (10-11) ◽  
pp. 731-735 ◽  
Author(s):  
K. Bourtzis ◽  
V. J. Marmaras
Keyword(s):  
Alkaline Phosphatase ◽  
Ceratitis Capitata ◽  
Divalent Cations ◽  
Deae Cellulose ◽  
Wild Type ◽  
Alkaline Phosphatases ◽  
Isolation And Characterization ◽  
Two Peaks ◽  
Ph Curve ◽  
Hydrolysis Of

Two specific alkaline phosphatase forms were identified in the integument of wild-type Ceratitis capitata during transition of larvae to pupae. The separation was achieved by DEAE-cellulose chromatography; alkaline phosphatase 1 and alkaline phosphatase 2 were eluted in 0.1 and 0.4 M KC1, respectively. Both isoenzymes have a molecular weight of approximately 180 000. The pH curve reveals two peaks for both alkaline phosphatases: one at 9.4 and the other at 11.0. The two isoenzymes at both pH optima catalyze the hydrolysis of phosphotyrosine and β-glycerophosphate, but not phosphoserine, phosphothreonine, ATP, or AMP. However, at pH 9.4, alkaline phosphatase 1 is more effective than ALPase 2 and exhibits a preference for phosphotyrosine. The divalent cations Mn2+, Mg2+, and Ba2+ activate the enzymes, while Cu2+ and Zn2+ are inhibitors for both isoenzymes. Both isoenzymes are inactivated by EDTA. The effect of amino acids on enzyme activity was also tested. Alkaline phosphatase 1 is inhibited by L-tyrosine, while alkaline phosphatase 2 is unaffected. L-Phenylalanine has no effect on either isoenzyme. Both isoenzymes are inhibited by urea and 2-mercaptoethanol. Simultaneous addition of urea and 2-mercaptoethanol reveals that ALPase 1 is more sensitive to these inhibitors than ALPase 2.Key words: enzyme, Diptera, integument, phosphatase, isoenzymes, phosphotyrosine.

scholarly journals Alkaline Phosphatase in Stem Cells

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Kateřina Štefková ◽  
Jiřina Procházková ◽  
Jiří Pacherník
Keyword(s):  
Stem Cells ◽  
Alkaline Phosphatase ◽  
Embryonic Stem ◽  
Stem Cell Marker ◽  
Developmental Studies ◽  
Alkaline Phosphatases ◽  
Potential Significance ◽  
Living Organisms ◽  
Almost All

Alkaline phosphatase is an enzyme commonly expressed in almost all living organisms. In humans and other mammals, determinations of the expression and activity of alkaline phosphatase have frequently been used for cell determination in developmental studies and/or within clinical trials. Alkaline phosphatase also seems to be one of the key markers in the identification of pluripotent embryonic stem as well as related cells. However, alkaline phosphatases exist in some isoenzymes and isoforms, which have tissue specific expressions and functions. Here, the role of alkaline phosphatase as a stem cell marker is discussed in detail. First, we briefly summarize contemporary knowledge of mammalian alkaline phosphatases in general. Second, we focus on the known facts of its role in and potential significance for the identification of stem cells.

scholarly journals The action of cyanate on human and pig kidney alkaline phosphatases

1969 ◽  
Vol 111 (5) ◽  
pp. 745-748 ◽  
Author(s):  
M. J. Carey ◽  
P. J. Butterworth
Keyword(s):  
Alkaline Phosphatase ◽  
Substrate Concentration ◽  
Ph Dependence ◽  
Thiol Group ◽  
Prolonged Treatment ◽  
Enzyme Molecule ◽  
Amino Groups ◽  
Alkaline Phosphatases ◽  
Pig Kidney ◽  
Substrate Concentrations

1. At concentrations of cyanate up to 0·2m there is an apparently reversible combination with alkaline phosphatase (EC 3.1.3.1), but higher concentrations inhibit alkaline phosphatase irreversibly by a process that is time-dependent. 2. The effect of 0·2m-cyanate on the enzymic reaction velocity depends on the substrate concentration. There is inhibition when the substrate concentration is 1·0mm or higher, but at lower substrate concentrations cyanate has an activating effect. 3. The pH-dependence of the reversible reaction suggests that cyanate may react with a thiol group at or near the active site of the enzyme, preventing a conformational change that is believed to be important in the mechanism of action of alkaline phosphatase. 4. Prolonged treatment with 0·6m-cyanate probably carbamoylates all free amino groups in the enzyme molecule and generates a new enzyme with decreased Vmax. and increased Km.

scholarly journals The glycoprotein nature of pig kidney diamine oxidase. Role of disulphide groups and arginine residues in the concanavalin A-diamine oxidase interaction

1988 ◽  
Vol 253 (1) ◽  
pp. 103-107 ◽  
Author(s):  
M A Shah ◽  
R Ali
Keyword(s):  
Concanavalin A ◽  
Diamine Oxidase ◽  
Con A ◽  
Acetylneuraminic Acid ◽  
Pig Kidney ◽  
Arginine Residues ◽  
Arginine Modification ◽  
Kinetics Of ◽  
Protein Moiety

Pig kidney diamine oxidase (DAO) was found to contain 5% (w/w) natural hexose, 3.25% glucosamine, 2.61% N-acetylglucosamine and 0.25% N-acetylneuraminic acid. The enzyme exhibited strong affinity towards concanavalin A (Con A) with a stoichiometry of 1:4.6. The kinetics of interaction approached an apparent first-order rate, with a rate constant (Kapp.) value of 1.5 × 10(-2) min-1. The enzyme reduced with dithiothreitol followed by alkylation with iodoacetamide showed an increase in the stoichiometry of the Con A-DAO interaction. Similarly arginine modification by phenylglyoxal caused decreased affinity, with an altered Kapp. value of 9.09 × 10(-3) min-1. The results suggest that, besides the carbohydrate content, the protein moiety of the enzyme also plays a significant role in the Con A-DAO interaction.

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