Tightly bound nucleotides of the energy-transducing ATPase, and their role in oxidative phosphorylation. I. The Paracoccus denitrificans system

Harris, David A.; John, P. S.; Radda, George K.

doi:10.1016/0005-2728(77)90053-6

Cited by 28 publications

(15 citation statements)

References 20 publications

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“…After detachment from membrane vesicles, the soluble F 1 -ATPase from P. denitrificans becomes functionally and structurally unstable (ref. 18 and unpublished results), thus hindering its isolation. In this work, these barriers were surpassed by tracking sulfite-activated ATPase activity (9,17) and by stabilizing the isolated F 1 and F 1 F O complexes of P. denitrificans in the presence of glycerol.…”

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confidence: 92%

“…The latter mechanism can be ruled out in P. denitrificans because no F 1 F O -ATPase activation is observed with disulfide reductants such as dithiothreitol (17). Instead, the presence of one or more intrinsic inhibitor proteins in the ATP synthase of P. denitrificans has been inferred from its activation as ATPase with limited trypsinolysis in a crude extract of an F 1 -ATPase of very low activity (18), as well as in the native F 1 F o -ATPase of inverted membrane vesicles (17).…”

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confidence: 99%

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A novel 11‐kDa inhibitory subunit in the F₁F_OATP synthase ofParacoccus denitrificansand related α‐proteobacteria

et al. 2009

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The F(1)F(O) and F(1)-ATPase complexes of Paracoccus denitrificans were isolated for the first time by ion exchange, gel filtration, and density gradient centrifugation into functional native preparations. The liposome-reconstituted holoenzyme preserves its tight coupling between F(1) and F(O) sectors, as evidenced by its high sensitivity to the F(O) inhibitors venturicidin and diciclohexylcarbodiimide. Comparison and N-terminal sequencing of the band profile in SDS-PAGE of the F(1) and F(1)F(O) preparations showed a novel 11-kDa protein in addition to the 5 canonical alpha, beta, gamma, delta, and epsilon subunits present in all known F(1)-ATPase complexes. BN-PAGE followed by 2D-SDS-PAGE confirmed the presence of this 11-kDa protein bound to the native F(1)F(O)-ATP synthase of P. denitrificans, as it was observed after being isolated. The recombinant 11 kDa and epsilon subunits of P. denitrificans were cloned, overexpressed, isolated, and reconstituted in particulate F(1)F(O) and soluble F(1)-ATPase complexes. The 11-kDa protein, but not the epsilon subunit, inhibited the F(1)F(O) and F(1)-ATPase activities of P. denitrificans. The 11-kDa protein was also found in Rhodobacter sphaeroides associated to its native F(1)F(O)-ATPase. Taken together, the data unveil a novel inhibitory mechanism exerted by this 11-kDa protein on the F(1)F(O)-ATPase nanomotor of P. denitrificans and closely related alpha-proteobacteria.

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A novel 11‐kDa inhibitory subunit in the F₁F_OATP synthase ofParacoccus denitrificansand related α‐proteobacteria

et al. 2009

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“…At low PMF, for example in environments with limited oxygen concentrations, this reaction can be reversed in several bacteria, which use the energy released from hydrolysis of ATP to maintain a PMF (von Ballmoos et al ., 2009). However, ATP synthases from several other bacteria display only very limited ATP hydrolysis activity, for example in Paracoccus denitrificans (Harris et al ., 1977), Bacillus subtilis (Hicks et al ., 1994), Thermus thermophilus (Nakano et al ., 2008) and Mycobacterium phlei (Higashi et al ., 1975).…”

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confidence: 99%

ATP synthase in slow- and fast-growing mycobacteria is active in ATP synthesis and blocked in ATP hydrolysis direction

Haagsma

Driessen

Hahn

et al. 2010

FEMS Microbiology Letters

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ATP synthase is a validated drug target for the treatment of tuberculosis, and ATP synthase inhibitors are promising candidate drugs for the treatment of infections caused by other slow-growing mycobacteria, such as Mycobacterium leprae and Mycobacterium ulcerans. ATP synthase is an essential enzyme in the energy metabolism of Mycobacterium tuberculosis; however, no biochemical data are available to characterize the role of ATP synthase in slow-growing mycobacterial strains. Here, we show that inverted membrane vesicles from the slow-growing model strain Mycobacterium bovis BCG are active in ATP synthesis, but ATP synthase displays no detectable ATP hydrolysis activity and does not set up a proton-motive force (PMF) using ATP as a substrate. Treatment with methanol as well as PMF activation unmasked the ATP hydrolysis activity, indicating that the intrinsic subunit ɛ and inhibitory ADP are responsible for the suppression of hydrolytic activity. These results suggest that the enzyme is needed for the synthesis of ATP, not for the maintenance of the PMF. For the development of new antimycobacterial drugs acting on ATP synthase, screening for ATP synthesis inhibitors, but not for ATP hydrolysis blockers, can be regarded as a promising strategy.

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“…Tightly coupled vesicles from Paracoccus denitrificans have been shown to catalyze only very low rates of ATP hydrolysis (29,30) while being capable of high rates of oxidative phosphorylation (31,32). The reason(s) why ATP synthase in P. denitrificans carries out apparently unidirectional catalysis is not known.…”

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Energy-dependent Transformation of F0·F1-ATPase in Paracoccus denitrificans Plasma Membranes

Zharova

Vinogradov

2004

Journal of Biological Chemistry

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F 0 ⅐F 1 -ATPases (ATP synthases) are the oligomeric molecular machines that couple ATP hydrolysis (synthesis) with proton translocation across the energy-transducing membranes in mitochondria, chloroplasts, and bacteria. The structural arrangement of the subunits within F 0 ⅐F 1 complexes of various organisms is assumed to be very similar (1-3). The hydrophilic F 1 is composed of a trimer of tightly packed ␣⅐␤-subunit pairs and one copy each of the ␦-, ⑀-, and ␥-subunits (Escherichia coli nomenclature for the subunits is used). Long rod-like ␥-subunit is asymmetrically positioned in the central cavity of the almost spherical globular ␣⅐␤ trimer. F 0 component is the hydrophobic complex composed of 10 -14 transmembraneously positioned c-, one a-, and two b-subunits. The hydrophilic parts of b-subunits are bound to F 1 (to one pair of ␣⅐␤-and one ␦-subunit), thus forming the peripheral stalk. The other central stalk is formed by ␥⅐⑀ complex, which interacts with c-subunit(s) arranged in a ring. F 1 bears three "catalytic" nucleotide-binding sites located on ␤-subunits, and F 0 serves as a proton-conducting path. It is generally believed that the coupling between the ATP hydrolysis (synthesis) and flow of protons across the membrane results from the consequence of the long distance conformational change: ␣⅐␤-pair-associated chemical catalysis 3 ␥⅐⑀ 3 ab 2 c n leading to rotation of the rotor (␥⅐⑀ bound to c-ring) within the stator (␣⅐␤ trimer fixed by two b-and one ␦-subunits) (4).The kinetics of ATP hydrolysis catalyzed by the soluble F 1 or membrane-bound F 0 ⅐F 1 preparations of the enzyme (coupled or uncoupled) are very complex (5). It has been documented that the key factor for such a complexity is a formation of so-called ADP(Mg 2ϩ )-inhibited form of the enzyme originally described (6) and kinetically characterized in a number of reports published by our (7-11) and other groups (12-16). Several phenomena such as hysteresis in onset of the catalytic activity (17), slow inhibition of ATPase by Mg 2ϩ (18), activation of ATP hydrolysis by sulfite and other anions (9,19,20), and the inhibitory effect of azide (9) can be consistently explained by the kinetic scheme in which the slowly reversible interconversion between catalytically competent enzyme-ADP intermediate and its inactive "isomer" plays the central role (5). Perhaps the most intriguing property of the ADP(Mg 2ϩ )-deactivated ATPase is that being inactive in ATP hydrolysis, it is fully competent in the ATP synthase activity (21,22).The ADP(Mg 2ϩ )/ATP-and possibly ⌬ H ϩ 1 -induced rearrangement within F 0 ⅐F 1 complex that trigger its ATP hydrolase and ATP synthase activities remains unclear. The ⑀-subunit, an endogenous inhibitor of the ATP hydrolase activity of F 1 and F 0 ⅐F 1 (23, 24), seems to be the most likely candidate for the triggering function. It has been shown that two distinct domains of the ⑀-subunit can exist in different conformations interacting differently with ␥-subunit (25-27). Most recently, it has been reported that the isolated ⑀...

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Tightly bound nucleotides of the energy-transducing ATPase, and their role in oxidative phosphorylation. I. The Paracoccus denitrificans system

Cited by 28 publications

References 20 publications

A novel 11‐kDa inhibitory subunit in the F₁F_OATP synthase ofParacoccus denitrificansand related α‐proteobacteria

A novel 11‐kDa inhibitory subunit in the F₁F_OATP synthase ofParacoccus denitrificansand related α‐proteobacteria

ATP synthase in slow- and fast-growing mycobacteria is active in ATP synthesis and blocked in ATP hydrolysis direction

Energy-dependent Transformation of F0·F1-ATPase in Paracoccus denitrificans Plasma Membranes

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Tightly bound nucleotides of the energy-transducing ATPase, and their role in oxidative phosphorylation. I. The Paracoccus denitrificans system

Cited by 28 publications

References 20 publications

A novel 11‐kDa inhibitory subunit in the F1FOATP synthase ofParacoccus denitrificansand related α‐proteobacteria

A novel 11‐kDa inhibitory subunit in the F1FOATP synthase ofParacoccus denitrificansand related α‐proteobacteria

ATP synthase in slow- and fast-growing mycobacteria is active in ATP synthesis and blocked in ATP hydrolysis direction

Energy-dependent Transformation of F0·F1-ATPase in Paracoccus denitrificans Plasma Membranes

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A novel 11‐kDa inhibitory subunit in the F₁F_OATP synthase ofParacoccus denitrificansand related α‐proteobacteria

A novel 11‐kDa inhibitory subunit in the F₁F_OATP synthase ofParacoccus denitrificansand related α‐proteobacteria