Identification of high molecular weight forms of inositol 1,4,5-trisphosphate 3-kinase in rat thymus and human lymphocytes

Abstract

A soluble inositol 1,4,5-triphosphate 3-kinase (Ins P 3-kinase) has been characterizes from extracts of rat thymus. The enzyme was shown to have a molecular weight within the range 98,000–114,000 M r as determined by regeneration of enzyme activity from sodium dodecyl sulphate polyacrylamide gels. The enzyme phosphorylates inositol 1,4,5-triphosphate (Ins P 3) to inositol 1,3,4,5-tetrakisphosphate (Ins P 4) with an apparent Km of 3.1 ± 0.4 μM. The enzyme is stimulated 4–6 fold by Ca 2+/calmodulin and is not recognised by polyclonal antisera raised against rat brain s P 3 3-kinase A. High levels of Ins P 3 3-kinase activity were also detected in soluble extracts of human lymphocyte preparations. The human lymphocyte enzyme was shown to have a molecular weight between 61,000 and 70,000 M r as judged by SDS-PAGE, and was stimulated approximately 10-fold in the presence of Ca 2+/calmodulin. These results establish that Ins P 3 3-kinase from rat thymus and human lymphocyte preparations represent high molecular weight isoenzymes of the Ins P 3 3-kinase family.