TY - JOUR
T1 - The inositol hexakisphosphate kinase family. Catalytic flexibility and function in yeast vacuole biogenesis
AU - Saiardi, A.
AU - Caffrey, J. J.
AU - Snyder, S. H.
AU - Shears, S. B.
PY - 2000/8/11
Y1 - 2000/8/11
N2 - Saiardi et al. (Saiardi, A., Erdjument-Bromage, H., Snowman, A., Tempst, P., and Snyder, S. H. (1999) Curr. Biol. 9, 1323-1326) previously described the cloning of a kinase from yeast and two kinases from mammals (types 1 and 2), which phosphorylate inositol hexakisphosphate (InsP6) to diphosphoinositol pentakisphosphate, a 'high energy' candidate regulator of cellular trafficking. We have now studied the significance of InsP6 kinase activity in Saccharomyces cerevisiae by disrupting the kinase gene. These ip6kΔ cells grew more slowly, their levels of diphosphoinositol polyphosphates were 60-80% lower than wild-type cells, and the cells contained abnormally small and fragmented vacuoles. Novel activities of the mammalian and yeast InsP6 kinases were identified; inositol pentakisphosphate (InsP5) was phosphorylated to diphosphoinositol tetrakisphosphate (PP-InsP4), which was further metabolized to a novel compound, tentatively identified as bisdiphosphoinositol trisphosphate. The latter is a new substrate for human diphosphoinositol polyphosphate phosphohydrolase. Kinetic parameters for the mammalian type 1 kinase indicate that InsP5 (K(m) = 1.2μM) and InsP6 (K(m) = 6.7 μM) compete for phosphorylation in vivo. This is the first time a PP-InsP4 synthase has been identified. The mammalian type 2 kinase and the yeast kinase are more specialized for the phosphorylation of InsP6. Synthesis of the diphosphorylated inositol phosphates is thus revealed to be more complex and interdependent than previously envisaged.
AB - Saiardi et al. (Saiardi, A., Erdjument-Bromage, H., Snowman, A., Tempst, P., and Snyder, S. H. (1999) Curr. Biol. 9, 1323-1326) previously described the cloning of a kinase from yeast and two kinases from mammals (types 1 and 2), which phosphorylate inositol hexakisphosphate (InsP6) to diphosphoinositol pentakisphosphate, a 'high energy' candidate regulator of cellular trafficking. We have now studied the significance of InsP6 kinase activity in Saccharomyces cerevisiae by disrupting the kinase gene. These ip6kΔ cells grew more slowly, their levels of diphosphoinositol polyphosphates were 60-80% lower than wild-type cells, and the cells contained abnormally small and fragmented vacuoles. Novel activities of the mammalian and yeast InsP6 kinases were identified; inositol pentakisphosphate (InsP5) was phosphorylated to diphosphoinositol tetrakisphosphate (PP-InsP4), which was further metabolized to a novel compound, tentatively identified as bisdiphosphoinositol trisphosphate. The latter is a new substrate for human diphosphoinositol polyphosphate phosphohydrolase. Kinetic parameters for the mammalian type 1 kinase indicate that InsP5 (K(m) = 1.2μM) and InsP6 (K(m) = 6.7 μM) compete for phosphorylation in vivo. This is the first time a PP-InsP4 synthase has been identified. The mammalian type 2 kinase and the yeast kinase are more specialized for the phosphorylation of InsP6. Synthesis of the diphosphorylated inositol phosphates is thus revealed to be more complex and interdependent than previously envisaged.
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U2 - 10.1074/jbc.M002750200
DO - 10.1074/jbc.M002750200
M3 - Article
C2 - 10827188
AN - SCOPUS:0034637548
SN - 0021-9258
VL - 275
SP - 24686
EP - 24692
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 32
ER -