CHARACTERIZATION OF A SMALL CHONDROITIN-SULFATE PROTEOGLYCAN ISOLATEDFROM THE MUCUS SURROUNDING THE EMBRYOS OF VIVIPARUS-ATER (MOLLUSCA, GASTROPODA)

A small chondroitin sulfate proteoglycan was isolated and partially ch aracterized for core protein and glycosaminoglycan structures from the mucus surrounding embryos in the developmental pouch of Viviparus ate r (Mollusca Gastropoda). The protein bearing polysaccharide nature was confirmed by gel-permeation chromatography separation of fractions po sitive to the uronic acid dosage, 7.5% SDS-PAGE under reducing conditi ons, sequential staining with alcian blue and ammoniacal silver. Its m olecular mass was calculated at about 228,800. After degradation of th e galactosaminoglycan components by chondroitinase ABC in the presence of proteinase inhibitors, the molecular mass of the core protein was determined at about 72,200. Treatment of the proteoglycan with keratan ase did not modify its electrophoretic migration. Isoelectric focusing of the core protein demonstrated a micro-heterogeneity with the prese nce of two isoforms with different isoelectric point, pI = 8.2 and 6.6 , in a ratio of about 1:2.2. The glycosaminoglycan component of the pr oteoglycan was characterized as chondroitin sulfate with a molecular m ass of about 30,750 composed of 5% non-sulfated unsaturated disacchari de, 94% monosulfated disaccharides (4-monosulfated to 6-monosulfated d isaccharide ratio of 1.36) and 1.5% disulfated disaccharides (in parti cular 1.3% 2,6-disulfated disaccharide) with a sulfate to carboxyl rat io of 0.96. Degradation of the chondroitin sulfate with chondroitinase ABC and ACII permitted to determine a percentage of glucuronic acid o f about 78.4. The proteoglycan isolated from the mucus surrounding the embryos of Viviparus ater is formed by a small core protein bearing a bout five chondroitin sulfate chains (80% chondroitin sulfate/20% derm atan sulfate) with potential function in the developmental processes o f molluscs embryos. (C) 1998 Elsevier Science B.V.