Purification and biochemical properties of a high-molecular-mass inositol 1,4,5-trisphosphate 3-kinase isoenzyme in human platelets.
The phosphorylation of inositol 1,4,5-trisphosphate (InsP3) to inositol 1,3,4,5-tetrakisphosphate (InsP4) is catalysed by InsP3 3-kinase. A method is presented for a rapid purification of the enzyme from human platelets. The purified enzyme was identified as a polypeptide of M(r) 69,000-70,000 after SDS/PAGE. It had a specific activity of 1.45 +/- 0.1 mumol/min per mg, and the degree of stimulation by Ca2+/calmodulin was 17-fold at saturating calmodulin and 10 microM free Ca2+. The Km for InsP3 and for ATP was 2.0 microM and 2.5 mM respectively. Human platelet InsP3 3-kinase was not recognized by immunodetection with anti-(InsP3 3-kinase A) or anti-(InsP3 3-kinase B) antibodies. These data provide the first biochemical evidence for the existence of a novel InsP3 3-kinase isoenzyme in human platelets, which is distinct from previously reported InsP3 3-kinase A and InsP3 3-kinase B.
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Institut de Recherche Interdisciplinaire en Biologie Humaine et Nucléaire (IRIBHN), Université Libre de Bruxelles, Belgium.
Vanweyenberg
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Erneux
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D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001792:Blood Platelets / Q000201:enzymology*; D001921:Brain / Q000201:enzymology; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007163:Immunosorbent Techniques; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D007527:Isoenzymes / Q000097:blood*; D008970:Molecular Weight; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000097:blood*; D051381:Rats
D002147:Calmodulin; D007527:Isoenzymes; D015544:Inositol 1,4,5-Trisphosphate; D000255:Adenosine Triphosphate; D017853:Phosphotransferases (Alcohol Group Acceptor); C051038:Inositol 1,4,5-trisphosphate 3-kinase; D002118:Calcium