Purification and properties of inositol-1,4-bisphosphate 4-phosphohydrolase from rat brain.
Inositol-1,4-bisphosphate 4-phosphohydrolase (inositol-1,4-bisphosphatase) was highly purified from a soluble fraction of rat brain. On SDS-polyacrylamide gel electrophoresis, the purified enzyme gave a single protein band and its molecular weight was estimated to be 42000. The isoelectric point of the enzyme was 4.3. The enzyme specifically hydrolyzed the 4-phosphomonoester linkage of inositol 1,4-bisphosphate. The Km value for inositol 1,4-bisphosphate was 30 microM, and it required Mg2+ for activity. Ca2+ was a competitive inhibitor with a Ki value of 60 microM as regards the Mg2+ binding. Li+, which is known to be a strong inhibitor of inositol 1-phosphatase (EC 3.1.3.25), inhibited the enzyme activity and caused 50% inhibition at a concentration of 1 mM (IC50 = 1 mM). Li+ was an uncompetitive inhibitor of substrate binding with a Ki value of 0.6 mM. These inhibitory parameters of Li+ were quite similar to those for inositol 1-phosphatase (IC50 = 1 mM, Ki = 0.3 mM). Thus, the effect of Li+ on decreasing the free inositol level with a subsequent decrease in agonist-sensitive phosphoinositides, is caused by its inhibition of multiple enzymes involved in conversion of inositol 1,4-bisphosphate to inositol.
D000818:Animals; D001921:Brain / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D002413:Cations, Divalent; D004591:Electrophoresis, Polyacrylamide Gel; D006019:Glycolysis; D007295:Inositol Phosphates / Q000378:metabolism; D007526:Isoelectric Point; D008094:Lithium / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D008970:Molecular Weight; D010744:Phosphoric Monoester Hydrolases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013379:Substrate Specificity
D002413:Cations, Divalent; D007295:Inositol Phosphates; C044259:inositol 1,4-bis(phosphate); D008094:Lithium; C053073:inositol-1,4-bisphosphate 4-phosphohydrolase; D010744:Phosphoric Monoester Hydrolases; D008274:Magnesium; D002118:Calcium