Identification and partial characterization of a lipophosphoglycan from a pathogenic strain of Entamoeba histolytica

An acidic glycoconjugate could be extracted from a delipidated residue fraction of [3H]galactose, [3H]mannose or [32P]orthophosphate metabolically labeled Entamoeba histolytica with water/ethanol/diethylether/pyridine/NH4OH (15:15:5:1:0.017). The radioactively labeled glycoconjugate comprised 50-55% of the total [3H]galactose label incorporated into macromolecules. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of the radiolabeled glycoconjugate showed two diffuse smears centering around 110 kDa and 45 kDa. Similar profiles were observed for both [3H]galactose- and [32P]orthophosphate-labeled glycoconjugate. No such bands were visible in [35S]methionine-labeled material. The hydrophobic nature of this glycoconjugate was inferred from its chromatographic behavior on phenyl-Sepharose. The molecule was rendered hydrophilic after digestion with phosphatidylinositol-specific phospholipase C. It was also sensitive to deamination by nitrous acid. Mild acid hydrolysis led to its fragmentation into smaller molecules as revealed by Sepharose 4B chromatography. Paper chromatographic analysis of the depolymerized [3H]galactose- and [3H]mannose-labeled fragments revealed that each was sensitive to alkaline phosphatase. The major dephosphorylated fragment migrated as an apparent galactose and mannose containing disaccharide which migrated identically to the Gal beta 1-4Man disaccharide derived from the lipophosphoglycan of Leishmania donovani. The above data support the existence of a major acidic glycoconjugate in E. histolytica bearing striking structural similarities to the lipophosphoglycan of Leishmania.