Electrochemical potential of protons in vesicles reconstituted from purified,proton-translocating adenosine triphosphatase |
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Authors: | Nobuhito Sone Masasuke Yoshida Hajime Hirata Harumasa Okamoto Yasuo Kagawa |
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Institution: | (1) Department of Biochemistry, Jichi Medical School, 329-04 Tochigi-Ken, Japan;(2) Present address: Department of Biochemistry, Institute of Brain Research, Tokyo University, Faculty of Medicine, Hongo, 113 Tokyo, Japan |
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Abstract: | Summary Measurements were made of the difference in the electrochemical potential of protons (
) across the membrane of vesicles reconstituted from the ATPase complex (TF
0
·F
1) purified from a thermophilic bacterium and P-lipids. Two fluorescent dyes, anilinonaphthalene sulfonate (ANS) and 9-aminoacridine (9AA) were used as probes for measuring the membrane potential (![Delta](/content/wq0752w67tv86u4p/xxlarge916.gif) ) and pH difference across the membrane ( pH), respectively.In the presence of Tris buffer the maximal ![Delta](/content/wq0752w67tv86u4p/xxlarge916.gif) and no pH were produced, while in the presence of the permeant anion NO
3
–
the maximal pH and a low ![Delta](/content/wq0752w67tv86u4p/xxlarge916.gif) were produced by the addition of ATP. When the ATP concentration was 0.24mm, the ![Delta](/content/wq0752w67tv86u4p/xxlarge916.gif) was 140–150 mV (positive inside) in Tris buffer, and the pH was 2.9–3.5 units (acidic inside) in the presence of NO
3
–
. Addition of a saturating amount of ATP produced somewhat larger ![Delta](/content/wq0752w67tv86u4p/xxlarge916.gif) and pH values, and the
attained was about 310 mV.By trapping pH indicators in the vesicles during their reconstitution it was found that the pH inside the vesicles was pH 4–5 during ATP hydrolysis.The effects of energy transfer inhibitors, uncouplers, ionophores, and permeant anions on these vesicles were studied. |
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Keywords: | |
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