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- PDB-8hgx: NMR solution structure of subunit epsilon of the Acinetobacter ba... -

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Basic information

Entry
Database: PDB / ID: 8hgx
TitleNMR solution structure of subunit epsilon of the Acinetobacter baumannii F-ATP synthase
ComponentsATP synthase epsilon chain
KeywordsELECTRON TRANSPORT / F-ATP synthase / subunit eosilon / bioenergetics / Acinetobacter / baumannii
Function / homology
Function and homology information


proton-transporting ATP synthase complex, catalytic core F(1) / proton-transporting ATP synthase activity, rotational mechanism / hydrolase activity / ATP binding / plasma membrane
Similarity search - Function
ATP synthase delta/epsilon subunit, C-terminal domain superfamily / ATP synthase, F1 complex, delta/epsilon subunit / ATP synthase, F1 complex, delta/epsilon subunit, N-terminal / F0F1 ATP synthase delta/epsilon subunit, N-terminal / ATP synthase, Delta/Epsilon chain, beta-sandwich domain
Similarity search - Domain/homology
ATP synthase epsilon chain
Similarity search - Component
Biological speciesAcinetobacter baumannii (bacteria)
MethodSOLUTION NMR / simulated annealing
AuthorsShin, J. / Grueber, G.
Funding support Singapore, 1items
OrganizationGrant numberCountry
National Research Foundation (NRF, Singapore) Singapore
CitationJournal: FASEB J / Year: 2023
Title: Atomic insights of an up and down conformation of the Acinetobacter baumannii F -ATPase subunit ε and deciphering the residues critical for ATP hydrolysis inhibition and ATP synthesis.
Authors: Wuan-Geok Saw / Khoa Cong Minh Le / Joon Shin / Jes Hui Min Kwek / Chui Fann Wong / Priya Ragunathan / Tuck Choy Fong / Volker Müller / Gerhard Grüber /
Abstract: The Acinetobacter baumannii F F -ATP synthase (α :β :γ:δ:ε:a:b :c ), which is essential for this strictly respiratory opportunistic human pathogen, is incapable of ATP-driven proton ...The Acinetobacter baumannii F F -ATP synthase (α :β :γ:δ:ε:a:b :c ), which is essential for this strictly respiratory opportunistic human pathogen, is incapable of ATP-driven proton translocation due to its latent ATPase activity. Here, we generated and purified the first recombinant A. baumannii F -ATPase (AbF -ATPase) composed of subunits α :β :γ:ε, showing latent ATP hydrolysis. A 3.0 Å cryo-electron microscopy structure visualizes the architecture and regulatory element of this enzyme, in which the C-terminal domain of subunit ε (Abε) is present in an extended position. An ε-free AbF -ɑβγ complex generated showed a 21.5-fold ATP hydrolysis increase, demonstrating that Abε is the major regulator of AbF -ATPase's latent ATP hydrolysis. The recombinant system enabled mutational studies of single amino acid substitutions within Abε or its interacting subunits β and γ, respectively, as well as C-terminal truncated mutants of Abε, providing a detailed picture of Abε's main element for the self-inhibition mechanism of ATP hydrolysis. Using a heterologous expression system, the importance of Abε's C-terminus in ATP synthesis of inverted membrane vesicles, including AbF F -ATP synthases, has been explored. In addition, we are presenting the first NMR solution structure of the compact form of Abε, revealing interaction of its N-terminal β-barrel and C-terminal ɑ-hairpin domain. A double mutant of Abε highlights critical residues for Abε's domain-domain formation which is important also for AbF -ATPase's stability. Abε does not bind MgATP, which is described to regulate the up and down movements in other bacterial counterparts. The data are compared to regulatory elements of F -ATPases in bacteria, chloroplasts, and mitochondria to prevent wasting of ATP.
History
DepositionNov 15, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Nov 22, 2023Provider: repository / Type: Initial release
Revision 1.1May 15, 2024Group: Database references / Category: database_2 / Item: _database_2.pdbx_DOI
Revision 1.2Jun 12, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: ATP synthase epsilon chain


Theoretical massNumber of molelcules
Total (without water)15,5121
Polymers15,5121
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551
NMR ensembles
DataCriteria
Number of conformers (submitted / calculated)21 / 200structures with the lowest energy
RepresentativeModel #1minimized average structure

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Components

#1: Protein ATP synthase epsilon chain / ATP synthase F1 sector epsilon subunit / F-ATPase epsilon subunit


Mass: 15511.765 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Acinetobacter baumannii (bacteria)
Gene: atpC, A7M90_08520, AB945B12_00682, Aba9201_12755, ABCAM1_0172, ABKPCSM17A_01727, ABR2091_0173, ABUW_3731, ACX61_17950, APC21_14385, APD31_00885, AUO97_06420, AYR68_18050, B7L45_18620, B9X95_ ...Gene: atpC, A7M90_08520, AB945B12_00682, Aba9201_12755, ABCAM1_0172, ABKPCSM17A_01727, ABR2091_0173, ABUW_3731, ACX61_17950, APC21_14385, APD31_00885, AUO97_06420, AYR68_18050, B7L45_18620, B9X95_01230, BAA1790NC_3496, BS065_18355, C2U32_15275, C6N18_19570, CBE85_14435, CBL15_17785, CSB70_3895, CTZ19_18500, D8O08_000335, DLI71_10775, DLI72_06180, DOL94_02920, DVA69_09710, E1A86_02075, E1A87_05110, E2532_15790, E2533_14640, E2534_11110, E2535_10530, E2536_13180, E2538_11270, E2539_11975, E2540_15325, E2541_09260, EA686_08570, EA706_05510, EA720_009765, EA722_10625, EGM95_19705, EKS29_01635, EWO96_15825, F2P40_12650, F4T85_15175, FDN00_02385, FE003_18665, FJU36_14065, FJU42_13255, FJU76_16610, FR761_02125, G3N53_14500, GNY86_14290, GSE42_00725, H0529_15450, H1058_00785, HB367_12610, HBK86_18985, HIN86_18905, IMO23_00750, NCTC13305_02274, NCTC13421_03737, SAMEA104305318_03328, SAMEA104305340_02247, SAMEA104305385_03000, SI89_14475
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: V5VHG0

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Experimental details

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Experiment

ExperimentMethod: SOLUTION NMR
NMR experiment
Conditions-IDExperiment-IDSolution-IDSample stateSpectrometer-IDType
111isotropic12D 1H-15N HSQC
222isotropic13D HN(CA)CB
232isotropic13D CBCA(CO)NH
242isotropic13D HNCA
252isotropic13D HN(CO)CA
161isotropic13D 1H-15N NOESY
373isotropic13D (H)CCH-TOCSY
383isotropic13D 1H-13C NOESY

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Sample preparation

Details
TypeSolution-IDContentsLabelSolvent system
solution10.5 mM [U-100% 15N] A. baumannii F-ATP synthase subunit epsilon, 20 mM sodium phosphate, 100 mM sodium chloride, 0.01 % w/v sodium azide, 90% H2O/10% D2O15N_sample90% H2O/10% D2O
solution20.5 mM [U-100% 15N] A. baumannii F-ATP synthase subunit epsilon, 20 mM sodium phosphate, 100 mM sodium chloride, 0.01 % w/v sodium azide, 90% H2O/10% D2O13C15N_sample90% H2O/10% D2O
solution30.5 mM [U-100% 15N] A. baumannii F-ATP synthase subunit epsilon, 20 mM sodium phosphate, 100 mM sodium chloride, 0.01 % w/v sodium azide, 100% D2O13C15N_sample100% D2O
Sample
Conc. (mg/ml)ComponentIsotopic labelingSolution-ID
0.5 mMA. baumannii F-ATP synthase subunit epsilon[U-100% 15N]1
20 mMsodium phosphatenatural abundance1
100 mMsodium chloridenatural abundance1
0.01 % w/vsodium azidenatural abundance1
0.5 mMA. baumannii F-ATP synthase subunit epsilon[U-100% 15N]2
20 mMsodium phosphatenatural abundance2
100 mMsodium chloridenatural abundance2
0.01 % w/vsodium azidenatural abundance2
0.5 mMA. baumannii F-ATP synthase subunit epsilon[U-100% 15N]3
20 mMsodium phosphatenatural abundance3
100 mMsodium chloridenatural abundance3
0.01 % w/vsodium azidenatural abundance3
Sample conditions
Conditions-IDIonic strengthLabelpHPressure (kPa)Temperature (K)
1100 mM15N_17 1 atm298 K
2100 mM13C15N_17 1 atm298 K
3100 mM13C15N_27 1 atm298 K

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NMR measurement

NMR spectrometerType: Bruker AVANCE / Manufacturer: Bruker / Model: AVANCE / Field strength: 700 MHz

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Processing

NMR software
NameDeveloperClassification
TopSpinBruker Biospincollection
NMRPipeDelaglio, Grzesiek, Vuister, Zhu, Pfeifer and Baxprocessing
NMRDrawDelaglio, Grzesiek, Vuister, Zhu, Pfeifer and Baxprocessing
SparkyGoddarddata analysis
CYANAGuntert, Mumenthaler and Wuthrichstructure calculation
CNSBrunger, Adams, Clore, Gros, Nilges and Readrefinement
RefinementMethod: simulated annealing / Software ordinal: 8
NMR representativeSelection criteria: minimized average structure
NMR ensembleConformer selection criteria: structures with the lowest energy
Conformers calculated total number: 200 / Conformers submitted total number: 21

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