Isolation of a phosphomonoesterase from human platelets that specifically hydrolyzes the 5-phosphate of inositol 1,4,5-trisphosphate.
Platelets, and a variety of other cells, rapidly hydrolyze the phosphoinositides in response to stimulation by agonists. One of the products of hydrolysis of phosphatidylinositol 4,5-diphosphate is inositol 1,4,5-trisphosphate, which recently has been suggested to mediate intracellular Ca2+ mobilization. We have found that human platelets contain an enzyme that degrades inositol 1,4,5-trisphosphate. We have isolated this soluble enzyme and find that it hydrolyzes the 5-phosphate of inositol 1,4,5-trisphosphate (Km = 30 microM, Vmax = 5.3 microM/min/mg of protein). The products of the reaction are inositol 1,4-diphosphate and phosphate. The apparent molecular weight of the enzyme is 38,000 as determined both by gel filtration and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the presence and absence of 2-mercaptoethanol. This enzyme is specific for inositol 1,4,5-trisphosphate. Other water soluble inositol phosphates as well as phosphorylated sugars are not hydrolyzed, while the only inositol containing phospholipid hydrolyzed is phosphatidylinositol 4,5-diphosphate at a rate less than 1% that for inositol 4,5-trisphosphate. The inositol 1,4,5-trisphosphate 5-phosphomonoesterase requires Mg2+ for activity and is inhibited by Ca2+, Ki = 70 microM. Li+, up to 40 mM, has no effect on enzyme activity. The duration and magnitude of any inositol 1,4,5-trisphosphate response in stimulated platelets may be determined by the activity of this enzyme.
Connolly
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Bross
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Majerus
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D001792:Blood Platelets / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate; D007295:Inositol Phosphates / Q000378:metabolism*; D000072181:Inositol Polyphosphate 5-Phosphatases; D007700:Kinetics; D008094:Lithium / Q000494:pharmacology; D008274:Magnesium / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D013379:Substrate Specificity; D013403:Sugar Phosphates / Q000378:metabolism*
D007295:Inositol Phosphates; D013403:Sugar Phosphates; D015544:Inositol 1,4,5-Trisphosphate; D008094:Lithium; D010744:Phosphoric Monoester Hydrolases; D000072181:Inositol Polyphosphate 5-Phosphatases; D008274:Magnesium; D002118:Calcium