Abstract
We demonstrate for the first time that functional electrogenic Na+-dependent amino acid transport is preserved for extended periods when purified brush border membranes prepared in hypotonic media are lyophilized and then rehydrated in buffer containing mannitol, NaSCN, and/or KSCN/valinomycin. Reconstituted lyophilized apical membranes from small intestine formed morphologically, physiologically, and thermodynamically normal vesicles which transportedl-alanine via system B into an osmotically active space energized by secondary active transport, as measured under equilibrium and nonequilibrium conditions. The lyophilized membranes are readily prepared and stored, thereby providing a means to pool large quantities of formed vesicles that are useful in examining cloned and reconstituted native amino acid transporter polypeptides.
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