Improvement of reconstitution of the Cl(-)-translocating ATPase isolated from Acetabularia acetabulum into liposomes and several anion pump characteristics.
The improved reconstitution of the Mono Q-III fraction, a Cl(-)-translocating ATPase, isolated from Acetabularia acetabulum (Ikeda et al. (1990) Biochemistry 29, 2057-2065) into liposomes rendered transport properties of this enzyme clear. The liposomes were prepared by the reversed-phase method using egg lecithin and cholesterol in a molar ratio of 2:1 and the purified ATPase was incorporated into the liposomes by a dialysis for 3 h. About 80% of the ATPase was incorporated into the liposomes. The weight ratio of the enzyme to lipid was 1:400-600. A sigmoid curve was obtained when the Cl(-)-transport activity of the enzyme was plotted against Cl- concentration. Hill's plot afforded a half-substrate concentration [S]0.5 of 45 mM and a Hill's coefficient n of 2.33. Effects of Br- and F- on the Cl(-)-transport were also examined in the reconstituted system, both halide ions decreased the 36Cl- efflux significantly. These kinetic data are in good agreement with the electrophysiological data presented by Tittor et al. ((1983) J. Membr. Biol. 75, 129-139).
Ohhashi
T
T
Faculty of Pharmaceutical Sciences, Okayama University, Japan.
Katsu
T
T
Ikeda
M
M
D000076:Acetabularia / Q000201:enzymology*; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D027321:Anion Transport Proteins; D000838:Anions; D001693:Biological Transport, Active; D002712:Chlorides / Q000378:metabolism*; D004566:Electrodes; D004591:Electrophoresis, Polyacrylamide Gel; D008081:Liposomes
D027321:Anion Transport Proteins; D000838:Anions; D002712:Chlorides; D008081:Liposomes; D000251:Adenosine Triphosphatases; C031652:anion-sensitive ATPases