Heterogeneity of bovine corneal proteokeratan sulfate

Proteokeratan sulfate was extracted and purified from bovine corneal stroma and then characterized by chemical and biochemical analyses. It was fractionated into several fractions by affinity chromatography on a concanavalin A-Sepharose column or by hydrophobic chromatography on a phenyl-Sepharose column. These fractions differed widely from one another in carbohydrate content, though no significant differences of their amino acid compositions were observed. One fraction (ca. 25%, on a dry weight basis) tightly bound to a concanavalin A-Sepharose column, compared with another fraction (ca. 65%) weakly bound to the same column, was poor in galactose and N-acetylglucosamine, but contained mannose in a high proportion. Fractions (ca. 30%) tightly bound to a phenyl-Sepharose column, in contrast to the one (ca. 66%) weakly bound, had low carbohydrate contents, like the fraction tightly bound to a concanavalin A-Sepharose column. Additionally, the fractions tightly bound to these affinity columns exhibited strong inhibitory actions on erythrocyte-concanavalin A agglutination. To obtain further details of the carbohydrate moiety of the proteokeratan sulfate, an attempt was made to separate and characterize peptidokeratan sulfate and Asn-linked oligosaccharide derived from some proteokeratan sulfate fractions. The present work revealed that the proteokeratan sulfate contains keratan sulfate and high mannose-type oligosaccharide in an approximate chain number ratio of 3.5:1.0, the keratan sulfate content varies widely and the oligosaccharide content increases with decrease of the keratan sulfate content, and the protein core is homogeneous at least with respect to the amino acid composition.