Reconstitution and partial purification of the glibenclamide-sensitive, ATP-dependent K+ channel from rat liver and beef heart mitochondria.
The transport properties of mitochondria are such that net potassium flux across the inner membrane determines mitochondrial volume. It has been known that K+ uptake is mediated by diffusive leak driven by the high electrical membrane potential maintained by redox-driven, electrogenic proton ejection and that regulated K+ efflux is mediated by an 82-kDa inner membrane K+/H+ antiporter. There is also long-standing suggestive evidence for the existence of an inner membrane protein designed to catalyze electrophoretic K+ uptake into mitochondria. We report reconstitution of a highly purified inner membrane protein fraction from rat liver and beef heart mitochondria that catalyzes electrophoretic K+ flux in liposomes and channel activity in planar lipid bilayers. The unit conductance of the channel at saturating [K+] is about 30 pS. Reconstituted K+ flux is inhibited with high affinity by ATP and ADP in the presence of divalent cations and by glibenclamide in the absence of divalent cations. The mitochondrial ATP-dependent K+ channel is selective for K+, with a Km of 32 mM, and does not transport Na+. K+ transport depends on voltage in a manner consistent with a channel activity that is not voltage-regulated. Thus, the mitochondrial ATP-dependent K+ channel exhibits properties that are remarkably similar to those of the ATP-dependent K+ channels of plasma membranes.
Paucek
P
P
Department of Pharmacology, Medical College of Ohio, Toledo 43699.
Mironova
G
G
Mahdi
F
F
Beavis
A D
AD
Woldegiorgis
G
G
Garlid
K D
KD
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002417:Cattle; D002848:Chromatography, DEAE-Cellulose; D004305:Dose-Response Relationship, Drug; D004553:Electric Conductivity; D004591:Electrophoresis, Polyacrylamide Gel; D005905:Glyburide / Q000494:pharmacology*; D008051:Lipid Bilayers; D008274:Magnesium / Q000494:pharmacology; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism; D008929:Mitochondria, Heart / Q000378:metabolism*; D008930:Mitochondria, Liver / Q000378:metabolism*; D008970:Molecular Weight; D011188:Potassium / Q000378:metabolism; D015221:Potassium Channels / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D051381:Rats; D013367:Submitochondrial Particles / Q000378:metabolism*
D008051:Lipid Bilayers; D008565:Membrane Proteins; D015221:Potassium Channels; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D008274:Magnesium; D011188:Potassium; D005905:Glyburide; D002118:Calcium