1990
DOI: 10.1042/bj2670441
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Solubilization and separation of inositol 1,3,4,5-tetrakisphosphate- and inositol 1,4,5-trisphosphate-binding proteins and metabolizing enzymes in rat brain

Abstract: The two inositol phosphate-binding proteins, the Ins(1,4,5)P3 (InsP3) and Ins(1,3,4,5)P4 (InsP4) receptors, and the two particulate InsP3-metabolizing enzymes, InsP3 5-phosphatase and InsP3 3-kinase, were solubilized with detergent from rat cerebellar membranes. These four activities are shown to be distinct molecular species by separation using a variety of protein chromatographic steps. The pharmacology of the partially purified InsP4-binding site indicates that the binding has a high affinity and selectivit… Show more

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Cited by 40 publications
(16 citation statements)
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“…More recently, evidence has accumulated favoring a biological role for inositol 1,3,4,5, Preparation of Membranes and Membrane Solubilization. Rat cerebellar membranes were prepared as described (26,27). Briefly, cerebella from 30 male Sprague-Dawley rats (200-300 g) were homogenized (Polytron setting 6, 8 s) at 0°C in 250 ml ofhomogenization buffer [50 mM Tris-HCl (pH 7.7), containing 1 mM EDTA, 1 mM 2-mercaptoethanol, 25 mg of phenylmethanesulfonyl fluoride, 1.2 mg of chymostatin, 1.2 mg of antipain, 1.2 mg of pepstatin, 2.4 mg of aprotinin, 2.4 mg of leupeptin, and 62.5 mg of N-carbobenzoxyphenylalanine].…”
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confidence: 99%
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“…More recently, evidence has accumulated favoring a biological role for inositol 1,3,4,5, Preparation of Membranes and Membrane Solubilization. Rat cerebellar membranes were prepared as described (26,27). Briefly, cerebella from 30 male Sprague-Dawley rats (200-300 g) were homogenized (Polytron setting 6, 8 s) at 0°C in 250 ml ofhomogenization buffer [50 mM Tris-HCl (pH 7.7), containing 1 mM EDTA, 1 mM 2-mercaptoethanol, 25 mg of phenylmethanesulfonyl fluoride, 1.2 mg of chymostatin, 1.2 mg of antipain, 1.2 mg of pepstatin, 2.4 mg of aprotinin, 2.4 mg of leupeptin, and 62.5 mg of N-carbobenzoxyphenylalanine].…”
mentioning
confidence: 99%
“…We (26,27) and others (28)(29)(30)(31) Preparation of Membranes and Membrane Solubilization. Rat cerebellar membranes were prepared as described (26,27).…”
mentioning
confidence: 99%
“…
A series of "P-labeled D-myo-inositol 1,3,4,5-tetrakisphosphate [Ins(l ,3,4,5)P41 analogues was enzymically prepared from the corresponding D-myo-inositol 1,4,5-trisphosphate [Ins( 1,4,5)P,] analogues using recombinant rat brain Ins(1 ,4,5)P3 3-kinase and [y-32P]ATP. Ins( l ,4,5)P, analogues with bulky groups at the 2-OH position, substitutions of phosphates by thiophosphates and D-6-deoxy-myo-Ins(l,4,5)P3 were tested.
…”
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confidence: 99%
“…Its role in calcium mobilization from non-mitochondria1 stores is now generally accepted. In mammalian cells, the Ins(1,4,5)P, signal can be terminated by a 5-phosphatase to yield D-myo-inositol 1,4-diphosphate [2, 31, and also by a 3,4,5)P4 is metabolized by the same 5-phosphatase that dephosphorylates Ins(1,4,5)P3, and also by a 3-phosphatase [14]. …”
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confidence: 99%
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