THE ASPARTIC PROTEINASE OF BARLEY IS A VACUOLAR ENZYME THAT PROCESSESPROBARLEY LECTIN IN-VITRO

Previous work suggested that the aspartic proteinase from Hordeum volg are (HvAP) would be a vacuolar protein in plant cells. Based on N-term inal sequencing we show that the in vitro-translated protein was trans located into the lumen of microsomal membranes, causing a concomitant removal of 25 amino acid residues from the protein. Vacuoles were puri fied from barley leaf protoplasts and were shown to contain all of the aspartic proteinase activity found in the protoplasts. This vacuolar localization of HvAP was confirmed with immunocytochemical electron mi croscopy using antibodies to HvAP in both barley leaf and root cells. In an attempt to discern a function for this protease, we investigated the ability of HvAP to process the C-terminal proregion of barley lec tin (BL) in vitro. Prolectin (proBL), expressed in bacteria, was proce ssed rapidly when HvAP was added. Using several means, we were able to determine that 13 amino acid residues at the C terminus of proBL were cleaved off, whereas the N terminus stayed intact during this incubat ion. Immunohistochemical electron microscopy showed that HvAP and BL a re co-localized in the root cells of developing embryos and germinatin g seedlings. Thus, we propose that the vacuolar HvAP participates in p rocessing the C terminus of BL.