DERMATAN SULFATE PROTEOGLYCANS FROM THE MINERALIZED MATRIX OF THE AVIAN EGGSHELL

The eggshell of the chicken is a useful model to study matrix componen ts which affect biomineralization. As an extension of our previous imm unohistochemical work which suggested the presence of dermatan sulfate proteoglycans in the mineralized region of the eggshell, a study was undertaken to characterize these molecules biochemically. After demine ralization with HCl and extraction with 4 M guanidinium chloride conta ining protease inhibitors, the extract was partitioned by anion exchan ge chromatography. Step elution with 0.25 M and 1.0 M sodium chloride resulted in the generation of two fractions, both of which contain cho ndroitinase-sensitive proteoglycans with molecular weights estimated a t 200,000 by gel electrophoresis, The proteoglycans in each fraction h ave core proteins with molecular weights of approximately 120,000 and glycosaminoglycans with average molecular weights of 22,000. Based on differential sensitivity to chondroitinase ABC and AC II, these glycos aminoglycans contain a small proportion of dermatan sulfate. The disac charide compositions of these glycosaminoglycans differ for the proteo glycans eluted with 0.25 M and 1.0 M sodium chloride. Those eluted wit h lower sodium chloride are enriched in unsulfated chondroitin and hav e much more 4-sulfated than 6-sulfated disaccharides; those eluted wit h 1.0 M sodium chloride contain primarily 4-sulfated disaccharides, a small amount of 6-sulfated disaccharides, and less unsulfated disaccha rides than the proteoglycans eluted with 0.25 M sodium chloride. The l arge difference in the proportions of unsulfated chondroitin may be th e reason for the elution at different sodium chloride concentrations. Both of the anion exchange column fractions contain other proteins in addition to the proteoglycans. These proteins are not separated from t he proteoglycans by a second anion exchange column or by molecular sie ve chromatography under dissociative conditions. Of particular interes t is the observation that the eggshell proteoglycans and their core pr oteins are recognized by a monoclonal antibody which recognizes an epi tope on the core protein of avian versican. This suggests that, in spi te of the large differences in the sizes of the core proteins of versi can and the eggshell proteoglycans, these core proteins share some hom ology, Because anionic molecules are thought to be important regulator s of biomineralization, and because preparations like those analyzed i n this study have been shown to influence in vitro calcium carbonate c rystallization, the eggshell proteoglycans may play a role in eggshell mineralization.