Cl- channels in basolateral renal medullary vesicles. VIII. Partial purification and functional reconstitution of basolateral mTAL Cl- channels.
Cl- channels fused from basolateral mTAL membranes into planar bilayers have distinctive functional characteristics which, when taken together, are unique among Cl- channels. The properties of these 50 to 60 pS channels can account for the characteristics of basolateral Cl- conductances in microperfused mTAL segments and thus may mediate net basolateral Cl- absorption in the intact mTAL. In the present studies, we solubilized basolateral membranes from rabbit mTAL. Since basolateral mTAL Cl- channels contain arginine- and lysine-rich domains, we exposed these solubilized membranes to sequential cation- and anion-exchange chromatography. The bound and unbound eluates from cation- and anion-exchange chromatography were reconstituted into proteoliposomes which, when fused into bilayers, yielded Cl- channels whose properties were virtually identical to those described above for native basolateral mTAL channels fused into bilayers. As judged by valinomycin-sensitive conductive 36Cl- uptake, proteoliposomes reconstituted from the unbound eluates after anion-exchange chromatography were enriched at least 30-fold in Cl- channel activity and had about 30% of the total Cl- channel activity solubilized in native vesicles.
Winters
C J
CJ
Department of Internal Medicine, University of Arkansas College of Medicine, Little Rock.
Reeves
W B
WB
Andreoli
T E
TE
D000818:Animals; D001485:Basement Membrane / Q000737:chemistry*; D018118:Chloride Channels / Q000737:chemistry / Q000302:isolation & purification* / Q000502:physiology; D002712:Chlorides / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D007679:Kidney Medulla / Q000737:chemistry*; D008051:Lipid Bilayers; D008564:Membrane Potentials; D011510:Proteolipids / Q000378:metabolism; D011817:Rabbits
D018118:Chloride Channels; D002712:Chlorides; D008051:Lipid Bilayers; D011510:Proteolipids; C020485:proteoliposomes