Isolation, purification and characterization of Cytosolic L-myo- Inositol-1-phosphate synthase from Marine macro Red Alga Sebdenia flabellata (J. Agardh) P.G. Parkinson
1*Dibyendu Sekhar Mahanty, 2Jukta Adhikari and 3Ankita Sarkar
1Plant Physiology, 2Plant Biochemistry and 3Plant Molecular Biology Laboratory Post Graduate Department of Botany, Barasat Government College, Kolkata-700124 (India) *Corresponding author: dibyendu.mahanty@gmail.com
ABSTRACT
Inositols are a large class of poly-hydroxylated cycloalkanes, also called cyclitols. Out of nine structural isomers of inositol, myoinositol is ubiquitously present in all organisms. It is a poly-hydroxy alcohol with six hydroxyl groups attached to each carbon atom of the hexose sugar which take part in metabolism namely signal transduction, cell membrane biogenesis, seed maturation, seed germination, growth, reproduction and as osmolytes to withstand abiotic stresses in plants. Marine water due to dissolved salts possesses low water potential which is a great challenge for normal plants for survival. Elevated level of myoinositol has been reported from the plants living in highly saline environment like marine aquatic ecosystems. Rise in salinity units in the sea water of the littoral zones has shown considerable increase in the production of this myo-inositol. The principal enzymes that catalyze the biosynthesis of myo-inositol are two, L-myo-inositol-1-phosphate synthase (MIPS; EC 5.5.1.4) an isomerase and L-myo-inositol-1- phosphate phosphatase (MIPP; EC 3.1.3.25) which has been detected, purified and characterized from a number of biological systems ranging from microbial to higher plants and animals. The present work deals with the isolation of the first enzyme MIPS and its characterization from marine red macro alga Sebdenia flabellata (J. Agardh) P.G. Parkinson growing naturally in the elevated salinity levels at Arabian Sea coast of Okha, Gujarat, India. L-myo-inositol-1-phosphate synthase (MIPS) from S. flabellata was partially purified using cold centrifugation, ammonium sulphate salting out (0-25% and 25-75% saturation) followed by DEAE cellulose column chromatography and Sephadex G-200 gel filtration. 6.46 fold purification of MIPS was achieved which were pooled for characterization experiments. MIPS was active in a temperature range of 30oC to 40oC and unstable in high temperature or prolonged room temperature, active in the pH range of 6.5 to 7.5 and had a sole substrate D-Glucose-6-phosphate, Vmax and Km values for the enzyme were determined. NH4 +, Mg2+, Na+, Ca2+ ions could promote and Pb2+, Cd2+, Hg2+ had inhibitory effect on the activity of the enzyme