Analytical characterization of beetroot vacuole membrane.

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Abstract
Vacuoles from beetroot (B. vulgaris L. var. esculenta Gurke) isolated by a mechanical procedure were osmotically lysed to separate the membrane and sap components for analysis. Approximately 62% of the vacuole proteins, 70% of the nondialyzable carbohydrates and almost all of the phospholipids and sterols were recovered in the membrane fraction. The vacuole membrane had a phospholipid:protein ratio of 0.68 and a sterol:phospholipid ratio of 0.21. Complex polar lipids (17) including phosphatides and glycolipids were tentatively identified. Phosphatidylcholine (54%) and phosphatidylethanolamine (24%) were the most prominent phosphoglycerides besides phosphatidylserine, phosphatidylglycerol, phosphatidylinositol and phosphatidic acid (1, 4, 5 and 12%, respectively). A putative sulfoglycoside and 2 major ceramide glycoside-like lipids, resembling those of animal lysosomes, were identified by TLC. High-resolution (sodium dodecyl sulfate) acrylamide gel electrophoresis of the polypeptides from the vacuole revealed 15 major bands with apparent MW ranging from 12,000-91,000. Selective elution experiments delineated those polypeptides that were peripheral membrane proteins or sap proteins adsorbed to the membrane, and those that exhibited hydrophobic interactions with the lipid core. Lectin labeling results indicated that most of the polypeptides from the membrane and from the sap were glycoproteins, probably, of the high-mannose type, characteristic of lysosomal enzymes that have undergone several stages of posttranslational modification.