Myo-inositol content in pteridophytes and the isolation and characterization of L-myo-inositol-1-phosphate synthase from Diplopterygium glaucum

Abstract

Myo-inositol is involved in normal growth and development of all living organisms and L-myo-inositol-1-phosphate synthase (MIPS; EC: 5.5.1.4) is responsible for its de novo synthesis. This enzyme has been reported for a number of life forms including plants, animals and bacteria. In the present study free myo-inositol has been detected in the common pteridophytes found in the Darjeeling Himalayas and the enzyme, L-myo-inositol-1-phosphate synthase has been partially purified from Diplopterygium glaucum (Thunb.) Nakai. A crude homogenate from the reproductive pinnules of D. glaucum was subjected to streptomycin sulphate precipitation and 0-70% ammonium sulphate fractionation followed by successive chromatography through DEAE-cellulose, Hexylagarose and BioGel A-0.5m columns. This resulted in a partial purification of the enzyme of about 81-fold with 13.5% recovery. The pteridophytic MIPS specifically utilized D-glucose-6-phosphte and NAD+ as its substrate and co-factor, respectively. It shows a pH optimum between 7.0 and 7.5 while the temperature maximum was 30°C. The enzyme activity was stimulated by NH4+, slightly inhibited by Na+, Ba2+ and Cd 2+, and strongly inhibited by Li+, Zn2+ and Hg2+. EDTA, pCMB and some substrate isomers like glucose-1-phosphate, fructose-6-phosphte and galactose-6-phosphate were inhibitory to the enzyme. The apparent molecular weight of the native D. glaucum MIPS was determined to be approximately 171 kDa.