Coupling factor 1 from Escherichia coli lacking subunits .delta. and .epsilon.: preparation and specific binding to depleted membranes, mediated by subunits .delta. or .epsilon.

Tuttas-Dörschug, R; Hanstein, Walter G.

doi:10.1021/bi00438a030

Cited by 19 publications

(3 citation statements)

References 41 publications

(56 reference statements)

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“…Previously, we had shown that the ␤Y331W mutation did not affect the enzymatic properties; ␤Y331W F 1 and F 1 F 0 are fully competent in ATP hydrolysis and synthesis (18,20). Here we first demonstrated that the interaction between ␤Y331W F 1 and the ⑀-subunit, with K d ϳ 1 nM, is very similar to that found in the wild-type enzyme (8,33) and that, as in wild-type F 1 (8), ⑀ reduced the ATPase activity of ␤Y331W F 1 by ϳ90%.…”

Section: Resultssupporting

confidence: 56%

Effect of the ε-Subunit on Nucleotide Binding toEscherichia coli F1-ATPase Catalytic Sites

Weber

Dunn

Senior

1999

Journal of Biological Chemistry

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The influence of the ⑀-subunit on the nucleotide binding affinities of the three catalytic sites of Escherichia coli F 1 -ATPase was investigated, using a genetically engineered Trp probe in the adenine-binding subdomain (␤-Trp-331). The interaction between ⑀ and F 1 was not affected by the mutation. K d for binding of ⑀ to ␤Y331W mutant F 1 was ϳ1 nM, and ⑀ inhibited ATPase activity by 90%. The only nucleotide binding affinities that showed significant differences in the ⑀-depleted and ⑀-replete forms of the enzyme were those for MgATP and MgADP at the high-affinity catalytic site 1. K d1 (MgATP) and K d1 (MgADP) were an order of magnitude higher in the absence of ⑀ than in its presence. In contrast, the binding affinities for MgATP and MgADP at sites 2 and 3 were similar in the ⑀-depleted and ⑀-replete enzymes, as were the affinities at all three sites for free ATP and ADP. Comparison of MgATP binding and hydrolysis parameters showed that in the presence as well as the absence of ⑀, K m equals K d3 . Thus, in both cases, all three catalytic binding sites have to be occupied to obtain rapid (V max ) MgATP hydrolysis rates.

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Section: Resultssupporting

confidence: 56%

Effect of the ε-Subunit on Nucleotide Binding toEscherichia coli F1-ATPase Catalytic Sites

Weber

Dunn

Senior

1999

Journal of Biological Chemistry

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“…However, it was recently determined that this efficiency factor was mainly due to the εNTD, at least for PS3-F 1 [92]. This observation with isolated F 1 does not rule out the additional role of the εCTD noted above for intact F O F 1 in membranes since, as noted earlier, the impact of ε on F 1 activity is clearly modulated by interactions of F 1 with F O (see also [93, 94]).…”

Section: Regulatory Elements Of Atp Synthases In Bacteria and Chlomentioning

confidence: 92%

The regulatory subunit ε in Escherichia coli FOF1-ATP synthase

Sielaff

Duncan

Börsch

2018

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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F-type ATP synthases are extraordinary multisubunit proteins that operate as nanomotors. The Escherichia coli (E. coli) enzyme uses the proton motive force (pmf) across the bacterial plasma membrane to drive rotation of the central rotor subunits within a stator subunit complex. Through this mechanical rotation, the rotor coordinates three nucleotide binding sites that sequentially catalyze the synthesis of ATP. Moreover, the enzyme can hydrolyze ATP to turn the rotor in the opposite direction and generate pmf. The direction of net catalysis, i.e. synthesis or hydrolysis of ATP, depends on the cell's bioenergetic conditions. Different control mechanisms have been found for ATP synthases in mitochondria, chloroplasts and bacteria. This review discusses the auto-inhibitory behavior of subunit ε found in FF-ATP synthases of many bacteria. We focus on E. coli FF-ATP synthase, with insights into the regulatory mechanism of subunit ε arising from structural and biochemical studies complemented by single-molecule microscopy experiments.

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“…ECF, was isolated from Escherichia coli strain AN 1460 by a modification of the method of Wise et al (1981) described in Gogol et al (1989a). ECF,, depleted of 5 and ( subunits (ECF,*), was prepared according to Tuttas-Dorschug and Hanstein (1989) using Sephacryl S-300 (Pharmacia) instead of Bio-Gel A 1.5M and also employing the same glycerol-containing buffers used in the ECF, purification. ECF,F0 was isolated from a sucrose gradient and reconstituted into egg lecithin as previously described (Aggeler et al, 1987).…”

Section: Methodsmentioning

confidence: 99%

Monoclonal antibody modification of the ATPase activity of Escherichia coli F1 ATPase

Aggeler¹,

Capaldi²

1990

Biochemistry

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Monoclonal antibodies (mAbs) have been made against each of the five subunits of ECF1 (alpha, beta, gamma, delta, and epsilon), and these have been used in topology studies and for examination of the role of individual subunits in the functioning of the enzyme. All of the mAbs obtained reacted with ECF1, while several failed to react with ECF1F0, including three mAbs against the gamma subunit (gamma II, gamma III, and gamma IV), one mAb against delta, and two mAbs against epsilon (epsilon I and epsilon II). These topology data are consistent with the gamma, delta, and epsilon subunits being located at the interface between the F1 and F0 parts of the complex. Two forms of ECF1 were used to study the effects of mAbs on the ATPase activity of the enzyme: ECF1 with the epsilon subunit tightly bound and acting to inhibit activity and ECF1* in which the delta and epsilon subunits had been removed by organic solvent treatment. ECF1* had an ATPase activity under standard conditions of 93 mumol of ATP hydrolyzed min-1 mg-1, cf. an activity of 7.5 units mg-1 for our standard ECF1 preparation and 64 units mg-1 for enzyme in which the epsilon subunit had been removed by trypsin treatment. The protease digestion of ECF1* reduced activity to 64 units mg-1 in a complicated process involving an inhibition of activity by cleavage of the alpha subunit, activation by cleavage of gamma, and inhibition with cleavage of the beta subunit. mAbs to the gamma subunit, gamma II and gamma III, activated ECF1 by 4.4- and 2.4-fold, respectively, by changing the affinity of the enzyme for the epsilon subunit, as evidenced by density gradient centrifugation experiments. The gamma-subunit mAbs did not alter the ATPase activity of ECF1*- or trypsin-treated enzyme. The alpha-subunit mAb (alpha I) activated ECF1 by a factor of 2.5-fold and ECF1F0 by 1.3-fold, but inhibited the ATPase activity of ECF1* by 30%.

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Coupling factor 1 from Escherichia coli lacking subunits .delta. and .epsilon.: preparation and specific binding to depleted membranes, mediated by subunits .delta. or .epsilon.

Cited by 19 publications

References 41 publications

Effect of the ε-Subunit on Nucleotide Binding toEscherichia coli F1-ATPase Catalytic Sites

Effect of the ε-Subunit on Nucleotide Binding toEscherichia coli F1-ATPase Catalytic Sites

The regulatory subunit ε in Escherichia coli FOF1-ATP synthase

Monoclonal antibody modification of the ATPase activity of Escherichia coli F1 ATPase

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