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

Schneider, Erwin; Altendorf, Karlheinz

doi:10.1002/j.1460-2075.1985.tb03658.x

Cited by 160 publications

(98 citation statements)

References 31 publications

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“…First, they may be important for anchoring the b subunits to a, maintaining the integrity of the stator complex. Second, given the observation that the b subunit is required for the formation of a functional proton pore upon reconstitution of F 0 from the purified subunits (47,48), the interactions between b and a in this region may have a role in assembly and organization of F 0 . This idea is further supported by evidence implicating the second stalk in the maturation of F 0 to a proton-conducting state (49,50).…”

Section: Figmentioning

confidence: 99%

Site-directed Cross-linking of b to the α, β, anda Subunits of the Escherichia coli ATP Synthase

McLachlin¹,

Coveny

Clark

et al. 2000

Journal of Biological Chemistry

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The b subunit dimer of the Escherichia coli ATP synthase, along with the ␦ subunit, is thought to act as a stator to hold the ␣ 3 ␤ 3 hexamer stationary relative to the a subunit as the ␥⑀c 9 -12 complex rotates. Despite their essential nature, the contacts between b and the ␣, ␤, and a subunits remain largely undefined. We have introduced cysteine residues individually at various positions within the wild type membrane-bound b subunit, or within b 24 -156 , a truncated, soluble version consisting only of the hydrophilic C-terminal domain. The introduced cysteine residues were modified with a photoactivatable cross-linking agent, and cross-linking to subunits of the F 1 sector or to complete ATP synthase, or F 1 F 0 -ATPase, utilizes a transmembrane proton gradient to synthesize ATP and is responsible for the final step in oxidative phosphorylation and photophosphorylation. The enzyme (reviewed in Refs. 1-3) is composed of two sectors. The membrane-integral F 0 sector is a proton pore, and in Escherichia coli has a subunit composition of ab 2 c 9 -12 . The membrane-peripheral F 1 sector has a subunit stoichiometry of ␣ 3 ␤ 3 ␥␦⑀. A key feature of the F 1 sector, as seen in the bovine heart mitochondrial crystal structure (4), is that the ␣ and ␤ subunits alternate in a ring around a lengthy pair of ␣-helices of ␥. Each ␤ subunit bears one catalytic nucleotide-binding site, while non-catalytic nucleotide-binding sites are found on the ␣ subunits. These nucleotide-binding sites are located close to the interfaces between ␣ and ␤ subunits, with one site near each of the six interfaces.Subunits from each sector contribute to the formation of two stalks that join F 1 and F 0 . The ␥ and ⑀ subunits form the central stalk, rotation of which is believed to be caused by translocation of protons across the membrane by the a and c subunits. This rotation is thought to cause conformational changes in the catalytic sites, driving synthesis of ATP (1). It is believed that the ␣ and ␤ subunits are prevented from rotating by a peripheral stalk consisting of ␦ and the two b subunits (5, 6) that joins the ␣ 3 ␤ 3 complex to the a subunit.In recent years the interaction of the b dimer and ␦ has been well established by a variety of evidence (7-10). The ␦ subunit appears to be located near the crown of the F 1 complex, the part of F 1 furthest from the membrane (11-15). Because b has a single membrane-spanning region at its N terminus, the remainder of the subunit must span a distance of over 100 Å to come in contact with ␦. Consistent with this proposed arrangement, the region of interaction between b and ␦ has been localized to the C terminus of b (10, 16). The hydrophilic domain of b by itself is mostly ␣-helical as measured by circular dichroism (17), and an isolated complex composed of ␦ with the hydrophilic domain of b was demonstrated by sedimentation velocity ultracentrifugation to be highly extended (9). The dimerization domain of b, encompassing residues 53-122, was also shown to be highly extended (18). Therefore the b 2...

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Section: Figmentioning

confidence: 99%

Site-directed Cross-linking of b to the α, β, anda Subunits of the Escherichia coli ATP Synthase

McLachlin¹,

Coveny

Clark

et al. 2000

Journal of Biological Chemistry

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“…Only when aJl three subunits were expressed was proton translocation detected. Schneider and Altendorf [5] dissociated isolated F 0 into its subunits. Only when all three subunits were reconstituted in stoichiometric amounts was proton translocation restored.…”

Section: Subunit Cmentioning

confidence: 99%

“…By removal of F 1 , the proton pathway in F 0 is opened, and can be studied independently [1][2][3][4]. Recently, a functional F 0 was reconstituted from its isolated components [5]. The availability of the complete primary structure of the eight subunits of the F 1 F 0 from E. coli and an increasing number of protein chemistry data, obtained with crosslinking reagents, proteolytic digestions and hydrophobic labeling techniques, has enabled us to propose models for the arrangement of the three F 0 subunits in the membrane [1][2][3][4][6][7][8][9][10][11][12][13][14][15][16][17][18].…”

Section: Introductionmentioning

confidence: 99%

Topological studies suggest that the pathway of the protons through F0 is provided by amino acid residues accessible from the lipid phase

Hoppe

Sebald

1986

Biochimie

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Summary -The structure of the F 0 part of ATP synthases from E. coli and Neurospora crassa was analyzed by hydrophobic surface labeling with [ 125 I]TID. In the E. co/i F 0 all three subunits were freely accessible to the reagent, suggesting that these subunits are independently integrated in the membrane. Labeted amino acid residues were identified by Edman degradation of the dicyclohexylcarbodiimide binding (DCCD) proteins from E. coli and Neurospora crassa. The very similar patterns obtained with the two homologaus proteins suggested the existence of tightly packed cx-helices. The oligomeric structure of the DCCD binding protein appeared to be very rigid since little, if any, change in the labeling patternwas observed upon addition of oligomycin or DCCD to membranes from Neurospora crassa. When membrancs were pretrcated with DCCD prior to the reaction with [1 25 I]TID an additionally labeled amino acid appeared at the position of Glu·65 which binds DCCD covalently, indicating the Jocation of this inhibitor on the outside of the oligomer. It is suggested that proton conduction occurs at the surface of the oligomer of the DCCD binding protein. Possibly this oligomer rotates against the subunit a or b and thus enables proton translocation. Conserved residues in subunit a, probably located in the Iipid bilayer, might participate in the pro· ton translocation mechanism.

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“…In this respect it is worthwhile mentioning that the i protein could also be solubilized in the absence of Fo subunits applying the purification protocols of Fo or FxFo to cells of the E. coli unc deletion strain CM 1470 (AuncI-A) transformed with plasmid pBS21 (data not shown). On the other hand, the presence of the i protein in samples of the Fo subunits a and b purified as described by Schneider and Altendorf [27] and its absence in any preparation of subunit c (data not shown) could be a hint for a possible association of the i protein with those proteins.…”

Section: 2 Detection and Localization Of The I Protehtmentioning

confidence: 79%

Detection and localization of thei protein inEscherichia coli cells using antibodies

1991

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

Cited by 160 publications

References 31 publications

Site-directed Cross-linking of b to the α, β, anda Subunits of the Escherichia coli ATP Synthase

Site-directed Cross-linking of b to the α, β, anda Subunits of the Escherichia coli ATP Synthase

Topological studies suggest that the pathway of the protons through F0 is provided by amino acid residues accessible from the lipid phase

Detection and localization of thei protein inEscherichia coli cells using antibodies

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