L-myo-Inositol 1-phosphate synthase from Entamoeba histolytica

Abstract

l-myo-Inositol 1-phosphate synthase (I-1-P synthase) catalyses the primary reaction for the synthesis of inositol in a variety of prokaryotes, eukaryotes and in the chloroplasts of algae and higher plants. Inositol is a precursor of essential macromolecules like membrane phospholipids, GPI anchor proteins and lipophosphoglycans, which play a determinant role in the pathogenesis of protozoan parasites such as Leishmania and Entamoeba. However, there is no report of I-1-P synthase or its gene from these organisms. The gene INO1 coding for this enzyme was first cloned from Saccharomyces cerevisiae and subsequently from several plants. Using molecular cloning techniques we have isolated and characterised the INO1 gene coding for the enzyme I-1-P synthase from Entamoeba histolytica. Simultaneously, we have purified and characterised the native enzyme from E. histolytica trophozoites and the cloned gene product from Escherichia coli. The gene product and the purified enzyme were both shown to be recognised by a heterologous anti-I-1-P synthase antibody from the phytoflagellate Euglena gracilis. Phylogenetic analysis of I-1-P synthase sequences from different eukaryotes suggest that it is highly conserved across species and the origin of this enzyme precedes the evolutionary divergence of modern eukaryotes.