EXPRESSION OF A NOVEL CELL-SURFACE LIPOPHOSPHOGLYCAN-LIKE GLYCOCONJUGATE IN TRYPANOSOMA-CRUZI EPIMASTIGOTES

The lipophosphoglycan (LPG)-like glycoconjugate expressed on the cell surface of Trypanosoma cruzi epimastigotes was isolated, purified, and partially characterized. The glycoconjugate migrated as a homogeneous band (42 kDa) on sodium dodecyl sulfate-polyacrylamide gel electropho resis. Matrix-assisted laser desorption ionization mass spectral analy sis of the native molecule indicated the presence of two major compone nts whose molecular masses were about 18.4 and 22.5 kDa. The LPG could be metabolically labeled with [H-3]galactose, [H-3]mannose, [C-14]glu cose, or [H-3]palmitic acid. Monosaccharide compositional analysis of the LPG indicated that galactose, glucosamine, and sialic acid predomi nate over mannose, galactosamine, and inositol. A peptide associated w ith the LPG molecule contained about 40 amino acid residues per inosit ol and had threonine as the predominant amino acid. The LPG showed str ong binding to Ricinus communis agglutinin-1 and Tritium vulgare wheat germ agglutinin, indicating the presence of terminal beta 1,4-linked galactosyl residue(s) and N-acetylglucosamine, respectively. Lectin bi nding studies also suggested the presence of a terminal beta-galactose and GlcNAc in the glycan-inositol lipid core of LPG. Virtually all of the sialic acids appeared to be located in the saccharide portion of the molecule. Treatment of the LPG with phosphatidylinositol-specific phospholipase C liberated an alkylacylglycerol. Structural analysis of the alkylacylglycerol and its acidic methanolysis products by gas-liq uid chromatography/ mass spectrometry indicated that the glycerol subs tituents were primarily the C-16 1-alkyl group and C-16 2-acyl group. The ratio of inositol to 1-O-alkyl-2-O-acylglycerol was 1:1. Treatment of the glycoconjugate with nitrous acid released a major phospholipid product that migrated close to the phosphatidylinositol standard on t hin layer chromatography. This result implied that phosphatidylinosito l was glycosidically linked to the nonacetylated amino sugar. Furtherm ore, the LPG was found to contain phosphate and was labile to mild aci d hydrolysis, strongly suggesting that the intact molecule is related to Leishmania LPG. The most striking and unique feature of T. cruzi LP G is the presence of large amounts of glucosamine and sialic acid as w ell as galactosamine. These results indicate that the glycoconjugate e xpressed on the T. cruzi cell surface is a new type of LPG-like molecu le anchored on the cell surface via an alkylacylphosphatidylinositol.