Purification, characterization, and localization of a 29 ku (Kilodalton) glycoprotein from the edible tubers of Apios americana Medikus

Total proteins from Apios americana Medikus tubers were resolved by SDS-PAGE into several abundant polypeptides of approximate molecular masses of 55, 42, 23, 22 and 14 ku (kDa). These proteins accumulated in a coordinate fashion during tuber development. We purified the 23 ku protein by ammonium sulfate precipitation, extraction with 70 % ethanol, and affinity chromatography on Con-A Sepharose. The amino acid composition of the purified 29 ku polypeptide revealed a low content of methionine and tyrosine. The N-terminal sequence of the purified 23 ku protein, EAKLPFFVFNLDRFF showed no significant homology to any known protein sequences. Mannose, glucose, galactose and N-Ac-glucosamine were detected by gas-liquid chromatography as the glycan components of the purified 29 ku glycoprotein. Antisera raised against the purified 29 ku glycoprotein contained antibodies directed against both the polypeptide moiety and the glycan side chains. Transmission electron microscopy immunocytochemical localization studies revealed that the 23 ku glycoprotein was found as discrete protein aggregates within vacuoles of Apios tuber cells.