A myo-inositol d-3 hydroxykinase activity in Dictyostelium

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Abstract
A soluble ATP-dependent enzyme which phosphorylates myo-inositol has been characterized in Dictyostelium. The myo-inositol kinase activity was partially purified from amoebae by chromatography on DEAE-Sepharose and phenyl-Sepharose columns. The product of both the partially purified activity and of a crude cytosolic fraction was myo-inositol 3-phosphate. The partially purified preparations of myo-inositol kinase (a) possessed a Km for myo-inositol of 120 .mu.M (in the presence of 5 mM-ATP) and for ATP of 125 .mu.M (in the presence of 1 .mu.M-myo-inositol), (b) did not recognize allo-, epi-, muco-, neo-, scyllo-, 1D-chiro or 1L-inositol as substrates, (c) were competitively inhibited by three naturally occurring analogues of myo-inositol; 1L-chiro-inositol (Ki 49.5 .+-. 0.7 .mu.M: the structural equivalent of myo-inositol, except that the D-3 hydroxy moiety is axial), D-3-deoxy-myo-inositol [Ki 103 .+-. 1 .mu.M: (-)-viburnitol], and sequoyitol (Ki 271 .+-. 7 .mu.M; unlike 1L-chiro-inositol and D-3-deoxy-myo-inositol, this was a substrate for the kinase), and finally (d) were apparently non-competitively inhibited by myo-inositol 3-phosphate. The product of myo-inositol kinase could be detected in intact amoebae and was a substrate for the first in a series of inositol polyphosphate kinase present in Dictyostelium which ultimately yield myo-inositol hexakisphosphate. The activity of myo-inositol D-3-hydroxykinase in Dictyostelium lysates showed evidence of developmental regulation.