Purification of bovine brain inositol‐1,4,5‐trisphosphate 5‐phosphatase

Verjans, Benoît; Lecocq, Raymond; Moreau, Colette; Erneux, Christophé

doi:10.1111/j.1432-1033.1992.tb16732.x

Cited by 29 publications

(20 citation statements)

References 24 publications

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“…Enzyme Preparation-Native bovine brain enzyme was purified as previously reported (27). The complete 412-amino acid-long coding region of human Type I Ins(1,4,5)P 3 5-phosphatase (plasmid ECH10 in Ref.…”

Section: Methodsmentioning

confidence: 99%

Arginine 343 and 350 Are Two Active Site Residues Involved in Substrate Binding by Human Type I D-myo-Inositol 1,4,5-Trisphosphate 5-Phosphatase

Communi¹,

Lecocq²,

Erneux³

1996

Journal of Biological Chemistry

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In a wide variety of cell types D-myo-inositol 1,4,5-trisphosphate (Ins(1,4,5)P 3 ) 1 and 1,2-diacylglycerol are generated from phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P 2 ) by receptor-mediated activation of phospholipase C (for review, see Refs. 1 and 2). Ins(1,4,5)P 3 mobilizes intracellular calcium from internal stores generating calcium signals to control many cellular processes: smooth muscle contraction, secretion, sensory perception, neuronal signaling, and cell growth (2). Ins(1,4,5)P 3 can be dephosphorylated by a 5-phosphatase to produce Ins(1,4)P 2 and phosphorylated by a 3-kinase to produce Ins(1,3,4,5)P 4 (3-5) (for review, see Refs. 2, 6, and 7). Some evidence supports a role for Ins(1,3,4,5)P 4 in the regulation of intracellular free calcium concentration in concert with Ins(1,4,5)P 3 (for review, see Refs. 7 and 8). Recently, a specific Ins(1,3,4,5)P 4 -binding protein has been isolated and identified as a member of the GAP1 family, suggesting a connection between phospholipase C-derived signals and a proliferative cascade involving Ras (9).A 75-kDa inositol polyphosphate 5-phosphatase was initially identified in human platelet lysates (10). cDNAs encoding the enzyme have been isolated from human cDNA libraries (11,12). When expressed in COS cells, it shows Ins(1,4,5)P 3 , Ins(1,3,4,5)P 4 , PtdIns(4,5)P 2 , and PtdIns(3,4,5)P 3 5-phosphatase activities (13,14). A protein identified due to its deficiency in the Lowe's oculocerebrorenal syndrome has been shown to be homologuous to the 75-kDa inositol polyphosphate 5-phosphatase (15). Expression of a truncated form of the protein demonstrates Ins(1,4,5)P 3 , Ins(1,3,4,5)P 4 , and PtdIns(4,5)P 2 5-phosphatase activities (13). In brain, Type I 43-kDa Ins(1,4,5)P 3 5-phosphatase is the major enzyme hydrolyzing the calcium-mobilizing second messenger Ins(1,4,5)P 3 . It hydrolyzes both Ins(1,4,5)P 3 and Ins(1,3,4,5)P 4 with higher affinity for Ins(1,3,4,5)P 4 but lower velocity (16). PtdIns(4,5)P 2 and PtdIns(3,4,5)P 3 are not substrates (14,17). cDNAs encoding Type I Ins(1,4,5)P 3 5-phosphatase have been isolated from several dog and human cDNAs libraries (18 -20).Arginyl residues are known to act as anionic binding sites in proteins and may thus assist in the binding of substrates or enzyme catalysis. Covalent and irreversible modification with amino acid specific reagents has been used successfully to identify lysyl or arginyl residues in the substrate binding domain in many enzymes, such as tyrocidine synthetase 1 (21), Ca 2ϩ /ATPase (22), 6-phosphofructo-2-kinase (23), and Ins(1,4,5)P 3 3-kinase (24). In addition, two arginyl residues were shown using site-directed mutagenesis to be critical to bind the C-2 phospho group of fructose 2,6-bisphosphate in rat * This work was supported by grants of the FRSM, Boehringer Ingelheim, and the Belgian Programme on Interuniversity Poles of Attraction initiated by the Belgian State, Prime Minister's Office, Federal Service for Science, Technology and Culture. The costs of publication of this article ...

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Section: Methodsmentioning

confidence: 99%

Arginine 343 and 350 Are Two Active Site Residues Involved in Substrate Binding by Human Type I D-myo-Inositol 1,4,5-Trisphosphate 5-Phosphatase

Communi¹,

Lecocq²,

Erneux³

1996

Journal of Biological Chemistry

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“…After washing with 100 mi of BD buffer, the enzyme was duted with a linear gradient of 0-1 M NaCI of 1 I (total volume). After concentration to 10 ml, further purification was achieved by Sephacryl S-200 gel filtration [5]: At this step, specific activity was 1.25/~mol/min/mg at 10 /zM InsP3. The pooled fractions were concentrated to 10 mi (1/zmol/ min/mi).…”

Section: Methodsmentioning

confidence: 99%

“…An afiquot of this preparation (0.25 ml) was six-fold diluted in 20 mM Na acetate, pH 5.5, and further purified onto a CM Mem Sep HP 1000 cation exchange HPLC resolved by a gradient of NaCI (0-0.2 M). Materials, assay of IusP3 5-phosphatase activity, Western blotting and immunodetection of proteins were as previously described [5][6].…”

Section: Methodsmentioning

confidence: 99%

Cloning and expression of human brain type I inositol 1,4,5‐trisphosphate 5‐phosphatase High levels of mRNA in cerebellar Purkinje cells

et al. 1994

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In brain and many other tissues, Type I inositol 1,4,5-trisphosphate (InsP3) 5-phosphatase is the major isozyme hydrolysing the calcium-mobilizing second messenger InsP3. We recently reported the cloning and expression of dog thyroid InsP3 5-phosphatase. During the course of this cloning, screening of a human brain eDNA library allowed us to isolate a eDNA clone D1 with 91% sequence identity with the thyroid sequence. When clone DI was expressed in Escherichia coli, the fusion protein had InsP3 5-phosphatase activity. Mr estimates of the recombinant enzyme made by immunodetection, activity assay after SDS/PAGE or silver staining were consistent with the calculated molecular mass. In situ hybridization on human cerebellum sections localised the mRNA for this enzyme to the Purkinje cells.

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“…5-Phosphatases lower IP 3 levels by dephosphorylating the 5Ј-phosphate, and they vary in substrate specificity Erneux et al, 1998). Type I 5-phosphatases are the most active in hydrolyzing IP 3 and IP 4 (Verjans et al, 1992;Laxminaryayan et al, 1993). Thus, they likely play a larger role in regulating cellular levels of IP 3 than do the type II phosphatases, which additionally hydrolyze the 5Ј-phosphoinositols, phosphatidyl inositol 4,5-bisphosphate and phosphatidyl inositol 3,4,5-trisphosphate .…”

Section: Introductionmentioning

confidence: 99%

Caenorhabditis elegansInositol 5-Phosphatase Homolog Negatively Regulates Inositol 1,4,5-Triphosphate Signaling in Ovulation

Bui

Sternberg

2002

MBoC

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Ovulation in Caenorhabditis elegans requires inositol 1,4,5-triphosphate (IP 3 ) signaling activated by the epidermal growth factor (EGF)-receptor homolog LET-23. We generated a deletion mutant of a type I 5-phosphatase, ipp-5, and found a novel ovulation phenotype whereby the spermatheca hyperextends to engulf two oocytes per ovulation cycle. The temporal and spatial expression of IPP-5 is consistent with its proposed inhibition of IP 3 signaling in the adult spermatheca. ipp-5 acts downstream of let-23, and interacts with let-23-mediated IP 3 signaling pathway genes. We infer that IPP-5 negatively regulates IP 3 signaling to ensure proper spermathecal contraction.

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Purification of bovine brain inositol‐1,4,5‐trisphosphate 5‐phosphatase

Cited by 29 publications

References 24 publications

Arginine 343 and 350 Are Two Active Site Residues Involved in Substrate Binding by Human Type I D-myo-Inositol 1,4,5-Trisphosphate 5-Phosphatase

Arginine 343 and 350 Are Two Active Site Residues Involved in Substrate Binding by Human Type I D-myo-Inositol 1,4,5-Trisphosphate 5-Phosphatase

Cloning and expression of human brain type I inositol 1,4,5‐trisphosphate 5‐phosphatase High levels of mRNA in cerebellar Purkinje cells

Caenorhabditis elegansInositol 5-Phosphatase Homolog Negatively Regulates Inositol 1,4,5-Triphosphate Signaling in Ovulation

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