Abstract

Cloned myo-inositol-1-phpsphate synthase (INOS) of Drosophila melanogaster was expressed in Escherichia coli, and purified using a His affinity column. The purified INOS required NAD^+ for the conversion of glucose-6-phosphate to inositol-1-phosphate. The optimum pH for myo-inositol-1-phosphate synthase is 7.5, and the maximum activity was measured at 40^oC. The molecular weight of the native enzyme, as determined by gel filtration, was approximately M_r 271,000±15,000. A single subunit of approximately M_r 62,000±5,000 was detected upon SDS-polyacrylamide gel electrophoresis. The Michaelis (K_m) and dissociation constants for glucose-6-phosphate were 3.5 and 3.7 mM, whereas for the cofactor NAD^+ these were 0.42 and 0.4 mM, respectively.

• Keywords: recombinant protein; inositol-1-phosphate synthase; multimeric protein; heat stability