scholarly journals Enzymic dephosphorylation of d-myo-inositol 1,4-bisphosphate in rat brain

1987 ◽  
Vol 242 (1) ◽  
pp. 193-198 ◽  
Author(s):  
A Delvaux ◽  
C Erneux ◽  
C Moreau ◽  
J E Dumont
Keyword(s):  
Rat Brain ◽  
Phosphatase Activity ◽  
Psychiatric Treatment ◽  
Soluble Fraction ◽  
Deae Cellulose ◽  
Brain Homogenate ◽  
Lithium Salts ◽  
Phosphatase Activities ◽  
Assay Mixture ◽  
Inositol 1,4,5 Trisphosphate

Inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] and inositol 1,4-bisphosphate [Ins(1,4)P2] phosphatase activities were measured in both 180,000 g (60 min) particulate and supernatant fractions of rat brain homogenates. Although Ins(1,4,5)P3 was mostly hydrolysed by a particulate phosphatase [Erneux, Delvaux, Moreau & Dumont (1986) Biochem. Biophys. Res. Commun. 134, 351-358], Ins(1,4)P2 phosphatase was predominantly soluble. The latter enzyme was Mg2+-dependent and sensitive to thiol-blocking agents (e.g. p-hydroxymercuribenzoate). In contrast with Ins(1,4,5)P3 phosphatase activity measured in the soluble fraction, Ins(1,4)P2 phosphatase was insensitive to 0.001-1 mM-2,3-bisphosphoglycerate. Lithium salts, widely used in psychiatric treatment, inhibited both Ins(1,4)P2 and Ins(1)P1 phosphatase activities of the crude soluble fraction. In particular, 50% inhibition of phosphatase activity, with 2 microM-Ins(1,4)P2 as substrate, was achieved at 3-5 mM-LiCl. At these concentrations, LiCl did not change Ins(1,4,5)P3 phosphatase activity measured in the same fraction with 1-4 microM-Ins(1,4,5)P3 as substrate. Chromatography of the soluble fraction of a rat brain homogenate on DEAE-cellulose resolved three phosphatase activities. These forms, peaks I, II and III, dephosphorylated Ins(1,4,5)P3, Ins(1)P1 and Ins(1,4)P2 respectively. If LiCl (10 mM) was included in the assay mixture, it inhibited both peak-II Ins(1)P1 phosphatase and peak-III Ins(1,4)P2 phosphatase, suggesting the existence of at least two Li+-sensitive phosphatases.

scholarly journals The dephosphorylation pathway of d-myo-inositol 1,3,4,5-tetrakisphosphate in rat brain

1987 ◽  
Vol 247 (3) ◽  
pp. 635-639 ◽  
Author(s):  
C Erneux ◽  
A Delvaux ◽  
C Moreau ◽  
J E Dumont
Keyword(s):  
Rat Brain ◽  
Soluble Fraction ◽  
Deae Cellulose ◽  
Phosphatase Activities ◽  
Inositol 1 ◽  
The Third ◽  
Inositol 1,4,5 Trisphosphate ◽  
Hydrolysis Of ◽  
Blue Sepharose

Dephosphorylation of inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P4] was measured in both the soluble and the particulate fractions of rat brain homogenates. Analysis of the hydrolysis of [4,5-32P]Ins(1,3,4,5)P4 showed that for both fractions the 5-phosphate of Ins(1,3,4,5)P4 was removed and inositol 1,3,4-trisphosphate [Ins(1,3,4)P3] was specifically produced. In the soluble fraction, Ins(1,3,4)P3 was further hydrolysed at the 1-phosphate position to inositol 3,4-bisphosphate[Ins(3,4)P2]. DEAE-cellulose chromatography of the soluble fraction separated the phosphatase activities into three peaks. The first hydrolysed both Ins(1,3,4,5)P4 and inositol 1,4,5-trisphosphate, the second inositol 1-phosphate and the third Ins(1,3,4)P3 and inositol 1,4-bisphosphate, [Ins(1,4)P2]. Further purification of the third peak on either Sephacryl S-200 or Blue Sepharose could not dissociate these two activities [i.e. with Ins(1,4)P2 and Ins(1,3,4)P3 as substrates]. The dephosphorylation of Ins(1,3,4)P3 could be inhibited by the addition of Li+.

scholarly journals The modulator protein dissociates the catalytic subunit of hepatic protein phosphatase G from glycogen

1988 ◽  
Vol 250 (3) ◽  
pp. 659-663 ◽  
Author(s):  
M Bollen ◽  
W Stalmans
Keyword(s):  
Phosphatase Activity ◽  
Rat Liver ◽  
Protein Phosphatase ◽  
Soluble Fraction ◽  
Glycogen Synthase ◽  
Gel Filtration ◽  
Catalytic Subunit ◽  
Regulatory Subunit ◽  
Phosphatase Activities ◽  
Glycogen Particles

1. The phosphorylase phosphatase and glycogen-synthase phosphatase activities associated with the glycogen particles from rat liver were progressively inhibited by incubation with modulator protein. However, the phosphorylase phosphatase activity of the catalytic subunit was entirely recovered after destruction of the modulator and the regulatory subunit(s) by trypsin. 2. Inhibition of protein phosphatase G by modulator was associated with a translocation of the phosphorylase phosphatase activity (measured after incubation with trypsin) from glycogen to the soluble fraction. The degree of inhibition of phosphatase G corresponded closely to the extent to which the phosphorylase phosphatase activity was released from the glycogen particles. Incubation of glycogen-free protein phosphatase G with modulator did not change the affinity of the enzyme for added glycogen, but decreased the amount of phosphatase that could be bound to glycogen. 3. The phosphorylase phosphatase activity that was released from the glycogen particles by modulator migrated on gel filtration as a complex (Mr 106,000) of the catalytic subunit with modulator. Phosphorylase phosphatase activity could be transferred from glycogen-bound protein phosphatase G to modulator that was covalently bound to Sepharose. After elution from the column, the enzyme was identified as the free catalytic subunit (Mr 37,000).

scholarly journals Synthesis of myo-inositol 1,3,4,5,6-pentakisphosphate from inositol phosphates generated by receptor activation

1988 ◽  
Vol 253 (3) ◽  
pp. 721-733 ◽  
Author(s):  
L R Stephens ◽  
P T Hawkins ◽  
C J Barker ◽  
C P Downes
Keyword(s):  
Rat Brain ◽  
Inositol Phosphates ◽  
Platelet Activating Factor ◽  
Chemical Properties ◽  
Brain Homogenate ◽  
Receptor Activation ◽  
Murine Bone Marrow ◽  
Astrocytoma Cells ◽  
Inositol 1,4,5 Trisphosphate ◽  
Stimulation Of

myo-[3H]Inositol 1,3,4,5,6-pentakisphosphate can be made from myo-[3H]inositol 1,4,5-trisphosphate in a rat brain homogenate or soluble fraction. Although D-myo-inositol 3,4,5,6-tetrakisphosphate can be phosphorylated by a soluble rat brain enzyme to give myo-inositol 1,3,4,5,6-pentakisphosphate, it is not an intermediate in the pathway from myo-inositol 1,4,5-trisphosphate. The intermediates in the above pathway are myo-inositol 1,3,4,5-tetrakisphosphate, myo-inositol 1,3,4-trisphosphate and myo-inositol 1,3,4,6-tetrakisphosphate [Shears, Parry, Tang, Irvine, Michell & Kirk (1987) Biochem. J. 246, 139-147; Balla, Guillemette, Baukal & Catt (1987) J. Biol. Chem. 262, 9952-9955], and it is catalysed by soluble kinase activities of similar anion-exchange mobility and Mr value. Compounds with chromatographic and chemical properties consistent with the structures myo-inositol 1,3,4,5-tetrakisphosphate, myo-inositol 1,3,4,6-tetrakisphosphate and myo-inositol 3,4,5,6-tetrakisphosphate are present in avian erythrocytes, human 1321 N1 astrocytoma cells and primary-cultured murine bone-marrow-derived macrophages. The amounts of these inositol tetrakisphosphates rise upon muscarinic cholinergic stimulation of the astrocytoma cells or stimulation of macrophages with platelet-activating factor.

Identification of the phosphomonoesterases that hydrolyze lysopolyphosphoinositides in rat brain and liver

1987 ◽  
Vol 65 (10) ◽  
pp. 890-898 ◽  
Author(s):  
Frederick B. St. C. Palmer
Keyword(s):  
Rat Brain ◽  
Phosphatase Activity ◽  
Divalent Cations ◽  
Liver Microsomes ◽  
Maximum Activity ◽  
Molar Ratio ◽  
Phosphatase Activities ◽  
Rat Brain And Liver ◽  
Ph Range

The phosphatase activities responsible for the sequential dephosphorylation of lysophosphatidylinositol 4,5-bisphosphate (lysoPtdIns(4,5)P2) to lysophosphatidylinositol that precedes reacylation in rat brain and liver microsomes were characterized. LysoPtdIns(4,5)P2 and the intermediate lysophosphatidylinositol 4-phosphate (lysoPtdIns4P) were hydrolyzed by two distinct phosphatase activities which were distinguishable by their substrate and cation requirements. The lysoPtdIns(4,5)P2 phosphatase activity was Mg2+ dependent and partially inhibited by Ca2+, excess Mg2+, and cationic detergent (cetyltrimethylammonium bromide). Activity was maximal at neutral (brain) or slightly alkaline (liver) pH when the Mg2+/lysoPtdIns(4,5)P2 molar ratio was 1.0 in the presence of bovine serum albumin (1 mg∙mL−1). LysoPtdIns4P phosphatase activity did not require divalent cations (not inhibited by EDTA). This activity was inhibited by Ca2+, Mg2+, and substrate concentrations above 0.2 mM. Maximum activity was observed over a broad pH range (6.0–8.5). Both activities were inhibited by lysophosphatidylinositol and lysophosphatidylcholine, but not other lysophospholipids. The lysopolyphosphoinositides are most likely hydrolyzed by the same phosphatases that act on the diacylpolyphosphoinositides, since PtdIns(4,5)P2 and PtdIns4P were also hydrolysed by Mg2+-dependent and cation-independent phosphatases, respectively. Activities with the diacylpolyphosphoinositides differed only in their requirement of detergents for maximum activity in vitro. Specific activities for the diacyl and "lyso" forms of each substrate were very similar when suitably optimized reaction mixtures were used. The subcellular distributions of the two phosphatase activities in both brain and liver were the same when acting on diacyl- or lyso-polyphosphoinositides, as was their response to inhibitors. Alkaline, acid, phosphoprotein, and inositol-1-phosphate phosphatases did not contribute substantially to the hydrolysis of either lysoPtdIns4P or lysoPtdIns(4,5)P2, since the activities were not significantly inhibited by cysteine, dithiothreitol, NaF, or LiCl. Lack of inhibition by 2,3-bisphosphoglycerate and absence of stimulation by cysteine or dithioerythritol, as well as a different subcellular distribution in liver, excluded inositol-1,4,5-trisphosphate and inositol-1,4-bisphosphate phosphatases as sources of the lysoPtdIns(4,5)P2 and lysoPtdIns4P phosphatase activities.

Ethanol and inositol 1,4,5-trisphosphate mobilize calcium from rat brain microsomes

Alcohol ◽  
1989 ◽  
Vol 6 (6) ◽  
pp. 431-436 ◽  
Author(s):  
Tina Machu ◽  
John J. Woodward ◽  
Steven W. Leslie
Keyword(s):  
Rat Brain ◽  
Inositol 1,4,5 Trisphosphate ◽  
Brain Microsomes

scholarly journals Formation of Monoenoic Fatty Acids by Desaturation in Rat Brain Homogenate

1973 ◽  
Vol 248 (5) ◽  
pp. 1786-1792
Author(s):  
Harold W. Cook ◽  
Matthew W. Spence
Keyword(s):  
Fatty Acids ◽  
Rat Brain ◽  
Brain Homogenate

scholarly journals Purification and properties of D-myo-inositol 1,4,5-trisphosphate 3-kinase from rat brain. Susceptibility to calpain.

1990 ◽  
Vol 265 (16) ◽  
pp. 9434-9440
Author(s):  
S Y Lee ◽  
S S Sim ◽  
J W Kim ◽  
K H Moon ◽  
J H Kim ◽  
...  
Keyword(s):  
Rat Brain ◽  
Purification And Properties ◽  
Inositol 1,4,5 Trisphosphate
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