Inhibition of porcine brain inositol 1,3,4-trisphosphate kinase by inositol polyphosphates, other polyol phosphates, polyanions and polycations.

We have partially purified an enzyme activity that phosphorylates inositol 1,3,4-trisphosphate from porcine brain, rat liver and bovine testis by FPLC chromatography on Q-Sepharose anion-exchange resin and Heparin-agarose. The products of this reaction were inositol 1,3,4,6-tetrakisphosphate and inositol 1,3,4,5-tetrakisphosphate. The same enzyme appears to be responsible for both 6-kinase and 5-kinase activities against inositol 1,3,4-trisphosphate (the 6-kinase: 5-kinase activity ratio is approximately 4 to 1), has a pH optimum of approximately 6.8 and requires Mg2+ for activity. The Km values of the enzyme for inositol 1,3,4-trisphosphate and ATP were approximately 0.5 microM and approximately 100 microM, respectively. Inositol 3,4,5,6-tetrakisphosphate, inositol 1,3,4,6-tetrakisphosphate and inositol 1,3,4,5-tetrakisphosphate are all competitive inhibitors with K(i) values of 0.4 microM, 3 microM and 5 microM, respectively, well within their likely intracellular concentration ranges: they inhibited 6-kinase and 5-kinase activities equally. 2,3-Bisphosphoglycerate and spermine were also competitive inhibitors, with K(i) values of 0.8 mM an 12 mM, respectively. Dextran sulphate was a non-competitive inhibitor with a Ki of approximately 15 microM, and poly-L-lysine (IC50 approximately 200 microM), polyvinylsulphate (IC50 approximately 250 microM) and heparin (IC50 approximately 2 mg/ml) also inhibited. Inhibition by these compounds suggests that inositol 3,4,5,6-tetrakisphosphate (and to a lesser extent inositol 1,3,4,5-tetrakisphosphate and other naturally occurring intracellular ions) may restrict the synthesis of inositol 1,3,4,6-tetrakisphosphate and hence regulate the rate of inositol penta- and hexakisphosphate synthesis from receptor-generated inositol phosphates.