Mechanism of Action of Mg2+ and Zn2+ on Rat Placental Alkaline Phosphatase. I. Studies on the Soluble Zn2+ and Mg2+ Alkaline Phosphatases

1975 ◽  
Vol 53 (10) ◽  
pp. 1089-1100 ◽  
Author(s):  
Claude PetitClerc ◽  
Monique Delisle ◽  
Marc Martel ◽  
Claude Fecteau ◽  
Normand Brière
Keyword(s):  
Alkaline Phosphatase ◽  
Catalytic Activity ◽  
Alkaline Phosphatase Activity ◽  
Active Sites ◽  
Placental Alkaline Phosphatase ◽  
Acidic Ph ◽  
Alkaline Ph ◽  
Enzyme Molecule ◽  
Alkaline Phosphatases ◽  
Enzyme Species

Rat placental alkaline phosphatase (EC 3.1.3.1), a dimer of 135 000 daltons, is strongly activated by Mg2+. However, Zn2+ has to be present on the apoenzyme to obtain this activation. Mg2+ alone is unable to reconstitute functional active sites. Excess Zn2+ which competes for the Mg2+ site leads to a phosphatase with little catalytic activity at alkaline pH, but with normal active sites at acidic pH as shown by covalent incorporation of ortho-[32P]phosphate.Two enzyme species with identical functional active sites have been reconstituted that only differ by the presence of Zn2+ or Mg2+ at the effector site.A mechanism is presented by which alkaline phosphatase activity of rat placenta would be controlled by a molecular process involving the interaction of Mg2+ and Zn2+ with the dimeric enzyme molecule.

Mechanism of action of Zn2+ and Mg2+ on rat placenta alkaline phosphatase. II. Studies on membrane-bound phosphatase in tissue sections and in whole placenta

1977 ◽  
Vol 55 (4) ◽  
pp. 474-478 ◽  
Author(s):  
Claude PetitClerc ◽  
Claude Fecteau
Keyword(s):  
Alkaline Phosphatase ◽  
Catalytic Activity ◽  
Active Sites ◽  
Maternal Serum ◽  
Alkaline Ph ◽  
Total Recovery ◽  
Tissue Sections ◽  
Trophoblastic Cells ◽  
Membrane Bound

Alkaline phosphatase (EC 3.1.3.1) bound to trophoblastic cells in rat placenta is activated by Mg2+ and inhibited by Zn2+ in the same way as is found with partially purified soluble alkaline phosphatase in the same tissue (PetitClerc, C., Delisle, M., Martel, M., Fecteau, C. &Brière, N. (1975) Can. J. Biochem. 53, 1089–1100). In studies done with tissue sections (6–10 μm), it is shown that alkaline phosphatase activity and labelling of active sites by orthophosphate are lost during incubation with ethanolamine at pH 9.0. Addition of Mg2+ causes total recovery of catalytic activity and active sites labelling. Zn2+ displaces and replaces at the Mg2+ binding sites. The affinity for both ions is similar, and dissociation of Zn2+ from the enzyme is a very slow process, even in the presence of Mg2+. The Zn2+–alkaline phosphatase and Mg2+–alkaline phosphatase, which only differ by the ion bound to an apparent modulator site, have the same catalytic activity at pH <7.0, but the Zn2+ species has little activity at alkaline pH. Phosphorylation of the enzyme by orthophosphate indicates that with both enzyme species phosphoryl intermediate does not accumulate at alkaline pH. These results suggest that with orthophosphate, the phosphorylation step is rate determining for both enzymes, and that Zn2+ affects this step to a much greater extent. It is proposed that Zn2+ and Mg2+ regulate alkaline phosphatase in rat placenta. The concentration of both ions in maternal serum and placenta suggest that such a mechanism could exist in vivo.

Macromolecular alkaline phosphatase and an immunoglobulin G that inhibited alkaline phosphatase in a patient's serum.

1983 ◽  
Vol 29 (2) ◽  
pp. 375-378 ◽  
Author(s):  
H Nakagawa ◽  
K Umeki ◽  
K Yamanaka ◽  
N Kida ◽  
S Ohtaki
Keyword(s):  
Alkaline Phosphatase ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Immunoglobulin G ◽  
Significant Loss ◽  
Placental Alkaline Phosphatase ◽  
Type Ii ◽  
Intestinal Alkaline Phosphatase ◽  
Adult Type ◽  
Heat Stable

Abstract Macromolecular alkaline phosphatase (EC 3.1.3.1) was found in the serum of a patient suffering from myasthenia gravis (adult type II) complicated with thymoma, and was shown by immunoelectrophoresis to be bound to immunoglobulins A and G (IgG). Placental alkaline phosphatase, complexed with either the patient's serum or IgG purified from the patient's serum, remained at the origin on electrophoresis, with significant loss of activity. Intestinal alkaline phosphatase, complexed with either the patient's serum or the patient's IgG, migrated to a position similar to that of the macromolecular alkaline phosphatase in the patient's serum on electrophoresis. About 50% of the placental alkaline phosphatase activity was inhibited with 0.1-0.2 g of the patient's IgG per liter, but 6.93 g of the IgG per liter was required for about 20% inhibition of the intestinal alkaline phosphatase activity. The complex of intestinal alkaline phosphatase with the patient's IgG was fairly heat stable. From these results, we concluded that the macromolecular alkaline phosphatase in the patient's serum consisted of intestinal alkaline phosphatase and IgG that was specific for placental alkaline phosphatase.

scholarly journals Alkaline Phosphatase Activity of Leukocytes in Shock

Blood ◽  
1955 ◽  
Vol 10 (7) ◽  
pp. 730-734 ◽  
Author(s):  
FRANCESCO VACCARI ◽  
BENITO SABOTTO ◽  
ENRICO MANZINI
Keyword(s):  
Alkaline Phosphatase ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Alkaline Phosphatases ◽  
General Metabolism

Abstract In animals and patients subjected to stress procedures and shock, there occurs simultaneously with the increase in general metabolism a rise of the cytoplasmic localizations of the alkaline phosphatases in granulocytes.

L-Phenylalanine inhibition of human alkaline phosphatases with p-nitrophenyl phosphate as substrate.

1982 ◽  
Vol 28 (12) ◽  
pp. 2426-2428 ◽  
Author(s):  
T Komoda ◽  
S Hokari ◽  
M Sonoda ◽  
Y Sakagishi ◽  
T Tamura
Keyword(s):  
Alkaline Phosphatase ◽  
Human Placenta ◽  
Placental Alkaline Phosphatase ◽  
Specific Inhibition ◽  
Human Intestine ◽  
Alkaline Phosphatases ◽  
Substrate Complex ◽  
Nitrophenyl Phosphate ◽  
Organ Specific ◽  
Alkaline Phosphatase Isoenzyme

Abstract With p-nitrophenyl phosphate as the substrate, there reportedly is no organ-specific inhibition of alkaline phosphatase (EC 3.1.3.1) activity by L-phenylalanine. However, we found that at pH 10.0, with p-nitrophenyl phosphate as the substrate, L-phenylalanine obviously inhibits the alkaline phosphatase isoenzyme from human placenta, whereas there is little if any inhibition of the isoenzyme from human intestine. Because of the differing effects of substrates (p-nitrophenyl phosphate and phenyl phosphate) and their enzymic products (p-nitrophenol and phenol) for L-phenylalanine action on the placental alkaline phosphatase isoenzyme, we suggest that the isoenzyme--inhibitor--substrate complex and the effect of released phosphate on L-phenylalanine inhibition of the isoenzyme activity differ from each other.

Influence of Light on the Alkaline Phosphatase Activity of Selenastrum capricornutum (Chlorophyceae) in Streams

1985 ◽  
Vol 42 (2) ◽  
pp. 384-388 ◽  
Author(s):  
R. L. Klotz
Keyword(s):  
New York ◽  
Alkaline Phosphatase ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
New York State ◽  
Selenastrum Capricornutum ◽  
Alkaline Phosphatases ◽  
Reactive Phosphorus ◽  
First Time

The alkaline phosphatase activity (APA) of Selenastrum capricomutum Printz incubated in situ in four streams in New York State was inversely related to total insolation. APA was not correlated with stream molybdate reactive phosphorus over the range of concentrations encountered. Selenastrum showed no diel cycle of APA. The phosphorus fraction made available by the activity of alkaline phosphatases, enzyme hydrolyzable phosphorus (measured for the first time in streams), increased the phosphorus supply to organisms with high APA.

scholarly journals Isolation and characterization of a Chinese hamster ovary (CHO) mutant defective in the second step of glycosylphosphatidylinositol biosynthesis

1996 ◽  
Vol 313 (1) ◽  
pp. 253-258 ◽  
Author(s):  
Victoria L. STEVENS ◽  
Hui ZHANG ◽  
Michelle HARREMAN
Keyword(s):  
Alkaline Phosphatase ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Second Step ◽  
Molecular Defect ◽  
Placental Alkaline Phosphatase ◽  
Isolation And Characterization

Mutant cell lines defective in the biosynthesis of glycosylphosphatidylinositol (GPI) described to date were isolated by selecting cells which no longer expressed one or more endogenous GPI-anchored proteins on their surface. In this study, a new mutant in this pathway was isolated from ethylmethanesulphonate-mutagenized Chinese hamster ovary cells stably transfected with human placental alkaline phosphatase (PLAP) as a marker of GPI-anchored proteins. A three-step protocol was employed. In the first step, cells with decreased surface expression of PLAP were selected by four rounds of complement-mediated lysis with an anti-(alkaline phosphatase) antibody. The surviving cells were cloned by limiting dilution and those with low levels of total alkaline phosphatase activity were selected in the second step. Finally, the ability of each clone to synthesize the first three intermediates in GPI biosynthesis in vitro was assessed to determine which cells with low alkaline phosphatase activity harboured a defect in one of these reactions. Of 230 potential mutants, one was defective in the second step of GPI biosynthesis. Microsomes from this mutant, designated G9PLAP.85, were completely unable to deacetylate either endogenous GlcNAc-phosphatidylinositol (PI) synthesized from UDP[6-3H]GlcNAc or exogenous GlcNAc-PI added directly to the membranes. Complementation analysis with the Thy-1-deficient murine lymphoma cells demonstrated that G9PLAP.85 has a molecular defect distinct from these previously described mutants. Therefore, these results suggest that mutants in GPI biosynthesis could be selected from almost any cell line expressing a GPI-anchored marker protein.

scholarly journals Gly429 is the major determinant of uncompetitive inhibition of human germ cell alkaline phosphatase by l-leucine

1991 ◽  
Vol 274 (1) ◽  
pp. 91-95 ◽  
Author(s):  
C Hummer ◽  
J L Millán
Keyword(s):  
Alkaline Phosphatase ◽  
Catalytic Activity ◽  
Germ Cell ◽  
Major Determinant ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Placental Alkaline Phosphatase ◽  
Uncompetitive Inhibition ◽  
Primary Determinant ◽  
Human Placental Alkaline Phosphatase

The catalytic activity of human placental alkaline phosphatase (PLAP) and germ cell alkaline phosphatase (GCAP) can be inhibited, through an uncompetitive mechanism, by L-Phe. GCAP is also selectively inhibited by L-Leu. Site-directed mutagenesis of five of the 12 residues which are different in PLAP and GCAP revealed that Gly429 is the primary determinant of GCAP inhibition by L-Leu, and Ser84 and Leu297 play a modulatory role in the inhibition.

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