Interaction of ovalbumin and its asparaginyl-carbohydrate fractions with concanavalin A.

The interaction of ovalbumin and its asparaginyl-carbohydrate fractions with concanavalin A was studied. Relative affinities were obtained by competitive binding studies using p-nitrophenyl alpha-D-mannopyranoside. Ovalbumin was separated into two fractions, I and II, by chromatography on concanavalin A-Sepharose. Ovalbumin and its fractions I and II interacted with concanavalin A in solution with binding affinities at 10 degrees C of 2 . 10(5) M-1, 3 . 10(4) M-1 and 2 . 10(6) M-1, respectively. The seven asparaginyl-carbohydrate fractions, obtained by fractionation on Dowex 50W-X2 (H+) and Durrum DA-4 (borate)columns, bound to concanavalin A with approximately the same affinity as native ovalbumin, suggesting that the sugar residues for binding in the isolated carbohydrates are exposed in the native protein. The binding of ovalbumin to concanavalin A was minimal after treatment with alpha-D-mannosidase in spite of the fact that only one half of the available mannose residues were hydrolyzed when compared to those removed by similar treatment of the asparaginyl-carbohydrate before fractionation. It is concluded that those alpha-D-mannosyl residues in ovalbumin that are required for binding to concanavalin A are accessible to alpha-D-mannosidase while the residual mannosyl groups are "buried" from interaction with concanavalin A and the enzyme.