Production of an Antibody Specific for the Propeptide of Wheat Germ Agglutinin

Wheat germ agglutinin (WGA) is synthesized as a proprotein with a glycosylated, 15 amino acid, carboxyl-terminal propeptide. This glycopeptide is cleaved from pro-WGA to produce the mature lectin during the transport of WGA to the protein bodies/vacuoles. To study the posttranslational modification of WGA, it would be useful to be able to differentiate between pro-WGA and mature WGA. Therefore, a peptide corresponding to the propeptide of WGA was synthesized (WGA-B 172-186), and an antiserum was raised in rabbits (anti-WGA-B 172-186). Anti-WGA-B 172-186 reacted with pure WGA-B 172-186 and pro-WGA in ELISA. Anti-WGA-B 172-186 was also specific for and readily differentiated between pro-WGA and mature WGA on Western blots. This provided an assay to monitor pro-WGA on Western blots before and after endo-beta-N-acetylglucosaminidase H digestion. Using this assay, direct evidence was obtained that the oligosaccharide of pro-WGA is of the high-mannose type.