C1^- channels in basolateral renal medullary vesicles VIII. Partial purification and functional reconstitution of basolateral mTAL C1^- 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.