THE PATHWAY OF MYOINOSITOL 1,3,4-TRISPHOSPHATE PHOSPHORYLATION IN LIVER - IDENTIFICATION OF MYOINOSITOL 1,3,4-TRISPHOSPHATE 6-KINASE, MYOINOSITOL 1,3,4-TRISPHOSPHATE 5-KINASE, AND MYOINOSITOL 1,3,4,6-TETRAKISPHOSPHATE 5-KINASE

  • 25 November 1989
    • journal article
    • research article
    • Vol. 264  (33) , 19879-19886
Abstract
Inositol 1,3,4-trisphosphate (Ins(1,3,4)P3) metabolism has been studied in liver homogenates and in 100,000 .times. g supernatant and particulate fractions. When liver homogenates were incubated in an "intracellular" medium containing 5 mM MgATP, equal proprotions of Ins(1,3,4)P3 were dephosphorylated and phosphorylated. Two inositol tetrakisphosphate (InsP4) products and an inositol pentakisphosphate (InsP5) were detected. The InsP4 isomerswere unequivocally identified as inositol 1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P4) and inositol 1,3,4,6-tetrakisphosphate (Ins(1,3,4,6)P4) by high performance liquid chromatography separation of inositol phosphates, periodate oxidation, alkaline hydrolysis, and stereospecific polyol dehydrogenase. Ins(1,3,4)P3 5-kinase is novel enzyme activity and accounted for 16% of the total Ins(1,3,4)P3 phosphorylation. Ins(1,3,46)P4 was also shown to be further phosphorylated to inositol 1,3,4,6-pentakisphosphate (Ins(1,3,4,5,6)P5) by a kinase not previously known to occur in liver. About 75% of Ins(1,3,4)P3 kinase activities were soluble and were partly purified by anion-exchange fast protein liquid chromatography. The two Ins(1,3,4)P3 kinase activities eluated as a single peak that was well resolved from Ins(1,3,4)P3 phosphatase, Ins(1,3,4,6)P4 5-kinase, and Ins(1,3,4,5)P4 5-phosphatase activities. A further novel observation was that 10 .mu.M Ins(1,3,4,5)P4 inhibited Ins(1,3,4)P3 kinase activites by 60%.