ATP‐synthetase complex (F1F0) from Escherichia coli

Steffens, Karl; Kiltz, Hans‐Hermann; Schneider, Erwin; Schmid, Roland; Altendorf, Karlheinz

doi:10.1016/0014-5793(82)80240-8

Cited by 11 publications

(6 citation statements)

References 22 publications

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“…Up till now, a study of the role of the single subunits of F0 in H+ translocation and F1 binding has been hampered by the fact that the subunits could be isolated in denatured form only (8).…”

Section: Discussionmentioning

confidence: 99%

“…Subunit b (Mr 17,265) is a rather hydrophilic protein, with only the NH2-terminal region (about 30 amino acid residues) buried within the membrane (12,13). By contrast, subunits a (Mr 30,276) and c (Mr 8,288) are very hydrophobic polypeptides. Whereas subunit a may span the membrane several times, subunit c is thought to form a hairpin-like structure within the membrane (14).…”

mentioning

confidence: 99%

“…When integrated into liposomes, purified Fo preparations exhibit passive H+ translocation and F1 binding (4,5). All subunits of Fo have been isolated and characterized under denaturing conditions (6)(7)(8). The DNA sequences of the corresponding genes have been established independently by three groups (9)(10)(11).…”

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confidence: 99%

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Subunit b of the membrane moiety (F0) of ATP synthase (F1F0) from Escherichia coli is indispensable for H+ translocation and binding of the water-soluble F1 moiety.

Schneider¹,

Altendorf²

1984

Proc. Natl. Acad. Sci. U.S.A.

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

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Subunit b of the membrane moiety (F0) of ATP synthase (F1F0) from Escherichia coli is indispensable for H+ translocation and binding of the water-soluble F1 moiety.

Schneider¹,

Altendorf²

1984

Proc. Natl. Acad. Sci. U.S.A.

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“…Fo [21] and subunits a, b and c [22] were purified under non-denaturing conditions from membranes of the ATP-synthase-overproducing strain KY7485 as described. Denatured subunits were prepared by different procedures : subunit c by chloroform/methanol extraction from whole cells of wild-type strain ML308-225 [32,331 and subunit b by BioGel P-30 gel filtration in the presence of 3% (w/v) SDS [34] using purified Fo as starting material. Subunit a was also prepared by the described gel filtration method, but for immunization further purification was necessary.…”

Section: Preparative Proceduresmentioning

confidence: 99%

“…Two sets of antisera have been prepared. Subunits isolated by chloroform/methanol extraction [33] or gel filtration in the presence of SDS [34] have been used as antigen (referred to as denatured subunits). In addition, all three Fo subunits have been purified under non-denaturing conditions as in [22].…”

Section: Immunoblottingmentioning

confidence: 99%

Accessibility of F₀ subunits from Escherichia coli ATP synthase

Deckers-Hebestreit

Altendorf

1986

European Journal of Biochemistry

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1. Antisera have been raised against denatured and non-denatured subunits a, b and c of the Fo complex of the ATP synthase from Escherichiu coli. The subunit specificity of the antibodies has been established with immunoblot analysis or enzyme-linked immunosorbent assay (ELISA).2. In inside-out oriented membrane vesicles the binding avidities of both sets of antisera, against denatured and non-denatured subunits of Fo, were similar in the presence as well as in the absence of the F1 part. F1-depleted everted membrane vesicles always produced an efficient binding of the different antisera. In the presence of F1 no antibody recognition could be observed with the anti-a antisera, while anti-b and anti-c antisera showed strong binding. However, a higher membrane protein concentration was necessary for the same antibody binding as in F1-stripped vesicles.3. In membrane vesicles with right-side-out orientation the recognition of the three Fo subunits was dependent on the antisera set used. Antisera raised against denatured subunits showed no binding to the membrane vesicles, only in case of anti-(dodecylsulfate-denatured b) antiserum could a slight affinity be detected. An antigen-antibody recognition with all three Fo subunits occurred when the antisera against non-denatured subunits were incubated with membrane vesicles of right-side-out orientation. The membrane protein concentration which was necessary to produce a significant binding was 10-100-fold higher compared to that of F,-depleted everted membrane vesicles.The ATP synthase (FIFO) of Escherichiu coli reversibly couples the synthesis or hydrolysis of ATP to the translocation of protons across the membrane. This enzyme consists of two structurally and functionally distinct entities: the watersoluble part (Fl), which carries the catalytic centers, and the membrane-integrated sector (Fo), which functions as a proton channel (for reviews, see [l -31). The Fo complex is composed of three different subunits a, b and c with relative molecular masses of 30276, 17265 and 8288, respectively and with the unusual stoichiometry of 1 : 2: 10 f 1 for a: b: c [4] (for reviews,Subunit c, also referred to as proteolipid or dicyclohexylcarbodiimide-binding protein, is the most extensively studied subunit of the Fo complex (for review, see [S] tyrosine residues [14] indicated that the hydrophilic turn is directed to the cytoplasmic side of the membrane. Antibody binding studies revealed that antigenic determinants of subunit c are accessible from the cytoplasm [15, 161. Subunit b has only a small hydrophobic segment at the N-terminal region with which it is possibly anchored into the membrane. The rest (80%) of the molecule is extremely hydrophilic [5, 61. This polar part is highly accessible to proteases and exposed to the cytoplasm Only a little is known about subunit a. Like subunit c, this protein is extremely hydrophobic spanning the membrane several times. The relative hydrophilic N-terminal region is probably exposed to the water phase [5, 61. Proteolytic degradation ...

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All three subunits are required for the reconstitution of an active proton channel (F0) of Escherichia coli ATP synthase (F1F0).

Schneider¹,

Altendorf²

1985

The EMBO Journal

160

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The membrane‐integrated, proton‐translocating F0 portion of the ATP synthase (F1F0) from Escherichia coli is built up from three kinds of subunits a, b and c with the proposed stoichiometry of 1:2:10 +/‐ 1. We have dissociated the F0 complex by treatment with trichloroacetate (3 M) at pH 8.0, in the presence of deoxycholate (1%) and N‐tetradecyl‐N, N‐dimethyl‐3‐ammonio‐1‐propanesulfonate (Zwittergent 3‐14, 5%). The subunits were separated by gel filtration with trichloroacetate (1 M) included in the elution buffer. The homogeneity of the fractions was checked by rechromatography and SDS‐gel electrophoresis. After integration into phospholipid vesicles each subunit alone as well as all possible combinations were tested for H+ translocating activity and binding of F1. A functional H+ channel could only be reconstituted by the combination a1b2c10 which corresponds to that of native F0.

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ATP‐synthetase complex (F₁F₀) from Escherichia coli

Cited by 11 publications

References 22 publications

Subunit b of the membrane moiety (F0) of ATP synthase (F1F0) from Escherichia coli is indispensable for H+ translocation and binding of the water-soluble F1 moiety.

Subunit b of the membrane moiety (F0) of ATP synthase (F1F0) from Escherichia coli is indispensable for H+ translocation and binding of the water-soluble F1 moiety.

Accessibility of F₀ subunits from Escherichia coli ATP synthase

All three subunits are required for the reconstitution of an active proton channel (F0) of Escherichia coli ATP synthase (F1F0).

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ATP‐synthetase complex (F1F0) from Escherichia coli

Cited by 11 publications

References 22 publications

Subunit b of the membrane moiety (F0) of ATP synthase (F1F0) from Escherichia coli is indispensable for H+ translocation and binding of the water-soluble F1 moiety.

Subunit b of the membrane moiety (F0) of ATP synthase (F1F0) from Escherichia coli is indispensable for H+ translocation and binding of the water-soluble F1 moiety.

Accessibility of F0 subunits from Escherichia coli ATP synthase

All three subunits are required for the reconstitution of an active proton channel (F0) of Escherichia coli ATP synthase (F1F0).

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ATP‐synthetase complex (F₁F₀) from Escherichia coli

Accessibility of F₀ subunits from Escherichia coli ATP synthase