Electrogenic K+ transport by the Kdp-ATPase of Escherichia coli.
Charge translocation by the Kdp-ATPase of Escherichia coli was measured by adsorption of proteoliposomes to a planar lipid membrane. The proteoliposomes were prepared by reconstitution of purified Kdp-ATPase into liposomes prepared from E. coli lipids. The protein was activated by a ATP concentration jump produced by photolysis of a protected derivative of ATP, caged ATP. Charge translocation was measured with a time resolution of 15-40 ms. Stationary currents demonstrated the continuous pumping activity of the enzyme. Control measurements with the potential-sensitive dye DiSC3(5) showed a negative potential inside the proteoliposomes after activation with ATP. The measured electrical signals as well as the dye measurements correspond to the transport of positive charge to the intracellular face of the protein. The electrical signal was increased when K+ was inside the proteoliposomes (K0.5 approximately 50 microM) and was inhibited by vanadate. These experiments demonstrate the electrogeneity of the Kdp-ATPase in a purified reconstituted system.
Fendler
K
K
Max-Planck-Institut für Biophysik, Frankfurt/M, Germany.
Dröse
S
S
Altendorf
K
K
Bamberg
E
E
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D052160:Benzothiazoles; D002232:Carbocyanines; D002352:Carrier Proteins / Q000378:metabolism*; D027682:Cation Transport Proteins; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D005456:Fluorescent Dyes; D007700:Kinetics; D008081:Liposomes; D009994:Osmolar Concentration; D011188:Potassium / Q000378:metabolism*; D011510:Proteolipids / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
D052160:Benzothiazoles; D002232:Carbocyanines; D002352:Carrier Proteins; D027682:Cation Transport Proteins; D029968:Escherichia coli Proteins; D005456:Fluorescent Dyes; D008081:Liposomes; D011510:Proteolipids; C020485:proteoliposomes; C012944:3,3'-dipropyl-2,2'-thiadicarbocyanine; C029273:P(3)-1-(2-nitro)phenylethyladenosine 5'-triphosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; C058166:potassium translocating Kdp-ATPase, E coli; D011188:Potassium