Binding of Concanavalin A to Milk Fat Globules and Release of the Lectin-Membrane Complex by Triton X-100

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Abstract
The interaction of concanavalin A with goat milk fat globules was investigated as 1 aspect of how this lectin suppresses milk secretion. These globules are coated with plasma membrane in their secretion from the cell and thus contain components of the lactating cell surface. Concanavalin A binding to globules was followed in fresh milk with the aid of [3H]concanavalin A, and membrane was released from lectin-treated and untreated globules with the nonionic detergent Triton X-100. Membrane pellets were obtained by centrifugation (50,000 .times. g, 1 h) and analyzed for protein, phospholipid [3H] activity, and peptide patterns by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Binding of the lectin to globules in milk at 37.degree. C was substantially complete in 30 min and fully reversed by 0.2 M .alpha.-methylmannoside. Of the concanavalin A added to milk, 30-40% associated with the fat globules; the rest was in various skim milk fractions. Binding curves revealed gradual saturation beginning about 12 mg of lectin bound per g of globules (1.3 mg of concanavalin A/mg of globule protein). The reaction of fat globules with concanavalin A in milk enhanced yields of membrane and may serve to stabilize globule structure during membrane preparation. Experiments employing [125I] and [125I] concanavalin A to label exposed proteins and concanavalin A receptors, respectively, provided evidence of a principal concanavalin A-binding glycoprotein (MW = 52,000) on the milk fat globule surface.