Sodium-dependent amino acid transport is preserved in lyophilized reconstituted apical membranes from intestinal epithelium.
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 transported L-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.
Stevens
B R
BR
Department of Physiology, College of Medicine, University of Florida, Gainesville 32610-0274, USA. stevens@phys.med.ufl.edu
Preston
R L
RL
D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D005612:Freeze Drying; D007413:Intestinal Mucosa / Q000378:metabolism* / Q000648:ultrastructure; D008297:Male; D008854:Microscopy, Electron; D008871:Microvilli / Q000378:metabolism / Q000648:ultrastructure; D011817:Rabbits; D012964:Sodium / Q000378:metabolism*
D000596:Amino Acids; D012964:Sodium