Aerobic Growth of Escherichia coli Is Reduced, and ATP Synthesis Is Selectively Inhibited when Five C-terminal Residues Are Deleted from the ϵ Subunit of ATP Synthase

Shah, Naman B.; Duncan, T. M.

doi:10.1074/jbc.m115.665059

Cited by 35 publications

(36 citation statements)

References 76 publications

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“…No ATP can be synthesized by ATP synthase carrying subunit a with the truncated N-terminus, while the wild type construct shows a rate of about 80 nmoles ATP/min/mg protein (Fig. 3), similar to other rates reported (Ishmukhametov et al 2008; Shah and Duncan 2015). Likewise, essentially no proton translocation can be seen from this mutant, using the energy of ATP hydrolysis (Fig.…”

Section: Resultssupporting

confidence: 87%

Analysis of an N-terminal deletion in subunit a of the Escherichia coli ATP synthase

Ishmukhametov

DeLeon-Rangel

Zhu

et al. 2017

J Bioenerg Biomembr

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Subunit a is a membrane-bound stator subunit of the ATP synthase and is essential for proton translocation. The N-terminus of subunit a in E. coli is localized to the periplasm, and contains a sequence motif that is conserved among some bacteria. Previous work has identified mutations in this region that impair enzyme activity. Here, an internal deletion was constructed in subunit a in which residues 6–20 were replaced by a single lysine residue, and this mutant was unable to grow on succinate minimal medium. Membrane vesicles prepared from this mutant lacked ATP synthesis and ATP-driven proton translocation, even though immunoblots showed a significant level of subunit a. Similar results were obtained after purification and reconstitution of the mutant ATP synthase into liposomes. The location of subunit a with respect to its neighboring subunits b and c was probed by introducing cysteine substitutions that were known to promote cross-linking: a_L207C + c_I55C, a_L121C + b_N4C, and a_T107C + b_V18C. The last pair was unable to form cross-links in the background of the deletion mutant. The results indicate that loss of the N-terminal region of subunit a does not generally disrupt its structure, but does alter interactions with subunit b.

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Section: Resultssupporting

confidence: 87%

Analysis of an N-terminal deletion in subunit a of the Escherichia coli ATP synthase

Ishmukhametov

DeLeon-Rangel

Zhu

et al. 2017

J Bioenerg Biomembr

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“…C), reflecting that W16 with its very high R2/R1 value and the hydrophobic patch of Mt ε plays a pivotal role in coupling the events of F 0 rotation with ATP synthesis via the εNTD to the εCTD and finally up to the DELSEED‐region of the catalytic α 3 :β 3 ‐domain. It is noteworthy that the deletion in the F‐ATP synthase mutant ε 1‐120 inhibited ATP synthesis by 65%, an effect which is similar to the truncation of the five C‐terminal ε‐hook residues of Ec ε, which significantly increased the inhibition of ATP synthesis and reduced the capacity for cell growth on a nonfermentable carbon source . However, an effect of the ε 1–120 mutant on M. smegmatis growth could not be observed in rich media (Fig.…”

Section: Discussionmentioning

confidence: 82%

The NMR solution structure of Mycobacterium tuberculosis F‐ATP synthase subunit ε provides new insight into energy coupling inside the rotary engine

et al. 2018

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“…One possibility that has been considered is that the e-subunit may contribute to this inhibitory mechanism as reported in other bacterial F-ATPases (11)(12)(13)(17)(18)(19)(20)(21). In the active F 1 -ATPase from G. stearothermophilus, for example, the e-subunit has been observed in the down position with the C-terminal α-helical hairpin, and bound ATP, alongside the N-terminal β-domain (17).…”

Section: Discussionmentioning

confidence: 95%

“…Up positions have been captured in structures of the F 1 -domains from E. coli (19,20), but the isolated e-subunit remains in a down conformation even when ATP is not bound to it (14)(15)(16). Deletion of its five C-terminal amino acids diminished respiratory growth (21), but deletion of the C-terminal domain had no growth phenotype (22).…”

mentioning

confidence: 99%

Regulation of the thermoalkaliphilic F ₁ -ATPase from Caldalkalibacillus thermarum

Ferguson

Cook

Montgomery

et al. 2016

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

107

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The crystal structure has been determined of the F 1 -catalytic domain of the F-ATPase from Caldalkalibacillus thermarum, which hydrolyzes adenosine triphosphate (ATP) poorly. It is very similar to those of active mitochondrial and bacterial F 1 -ATPases. In the F-ATPase from Geobacillus stearothermophilus, conformational changes in the e-subunit are influenced by intracellular ATP concentration and membrane potential. When ATP is plentiful, the e-subunit assumes a "down" state, with an ATP molecule bound to its two C-terminal α-helices; when ATP is scarce, the α-helices are proposed to inhibit ATP hydrolysis by assuming an "up" state, where the α-helices, devoid of ATP, enter the α 3 β 3 -catalytic region. However, in the Escherichia coli enzyme, there is no evidence that such ATP binding to the e-subunit is mechanistically important for modulating the enzyme's hydrolytic activity. In the structure of the F 1 -ATPase from C. thermarum, ATP and a magnesium ion are bound to the α-helices in the down state. In a form with a mutated e-subunit unable to bind ATP, the enzyme remains inactive and the e-subunit is down. Therefore, neither the γ-subunit nor the regulatory ATP bound to the e-subunit is involved in the inhibitory mechanism of this particular enzyme. The structure of the α 3 β 3 -catalytic domain is likewise closely similar to those of active F 1 -ATPases. However, although the β E -catalytic site is in the usual "open" conformation, it is occupied by the unique combination of an ADP molecule with no magnesium ion and a phosphate ion. These bound hydrolytic products are likely to be the basis of inhibition of ATP hydrolysis.

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Aerobic Growth of Escherichia coli Is Reduced, and ATP Synthesis Is Selectively Inhibited when Five C-terminal Residues Are Deleted from the ϵ Subunit of ATP Synthase

Cited by 35 publications

References 76 publications

Analysis of an N-terminal deletion in subunit a of the Escherichia coli ATP synthase

Analysis of an N-terminal deletion in subunit a of the Escherichia coli ATP synthase

The NMR solution structure of Mycobacterium tuberculosis F‐ATP synthase subunit ε provides new insight into energy coupling inside the rotary engine

Regulation of the thermoalkaliphilic F ₁ -ATPase from Caldalkalibacillus thermarum

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Aerobic Growth of Escherichia coli Is Reduced, and ATP Synthesis Is Selectively Inhibited when Five C-terminal Residues Are Deleted from the ϵ Subunit of ATP Synthase

Cited by 35 publications

References 76 publications

Analysis of an N-terminal deletion in subunit a of the Escherichia coli ATP synthase

Analysis of an N-terminal deletion in subunit a of the Escherichia coli ATP synthase

The NMR solution structure of Mycobacterium tuberculosis F‐ATP synthase subunit ε provides new insight into energy coupling inside the rotary engine

Regulation of the thermoalkaliphilic F 1 -ATPase from Caldalkalibacillus thermarum

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Regulation of the thermoalkaliphilic F ₁ -ATPase from Caldalkalibacillus thermarum