Mild alkaline borohydride treatment of glycoproteins—A method for liberating both N- and O-linked carbohydrate chains

Abstract

The generally accepted concept that N-linked oligosaccharides in glycoproteins are considerably more stable to mild alkali than are O-linked chains has been reexamined. A number of 3H-labeled model glycoproteins (fetuin, transferrin, and glycophorin) were treated with: (i) 0.05 m OH −−1 m BH 4 − at 50°C for 16 h, (ii) 0.1 m OH −−0.8 m BH 4 at 37°C for 68 h, and (iii) 1 m OH −−4 m BH 4 − at 80°C for 24 h. Analysis of the products by gel filtration and paper electrophoresis showed that both N- and O-linked chains were released by all three conditions. A portion (40%) of the asparagine-linked units produced under condition (i) remained as glycopeptides. Although mild alkaline borohydride cannot be used as a diagnostic reagent for distinguishing N- and O-glycosides, it is a useful general procedure for the analysis of the carbohydrate moieties of glycoproteins. Application of the method to a differentiation antigen (gp 160) of human kidney epithelia is described.