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- PDB-8k16: KtrA bound with ATP and thallium -

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Basic information

Entry
Database: PDB / ID: 8k16
TitleKtrA bound with ATP and thallium
ComponentsKtr system potassium uptake protein A
KeywordsMETAL TRANSPORT / potassium channel RCK domain
Function / homology
Function and homology information


monoatomic cation transmembrane transporter activity / potassium ion transport / identical protein binding / plasma membrane
Similarity search - Function
: / TrkA-N domain / Regulator of K+ conductance, C-terminal / Regulator of K+ conductance, C-terminal domain superfamily / TrkA-C domain / RCK C-terminal domain profile. / Regulator of K+ conductance, N-terminal / RCK N-terminal domain profile. / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / THALLIUM (I) ION / Ktr system potassium uptake protein A
Similarity search - Component
Biological speciesBacillus subtilis (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.1 Å
AuthorsChiang, W.T. / Chang, Y.K. / Hu, N.J. / Tsai, M.D.
Funding support Taiwan, 1items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, Taiwan) Taiwan
CitationJournal: Nat Commun / Year: 2024
Title: Structural basis and synergism of ATP and Na activation in bacterial K uptake system KtrAB.
Authors: Wesley Tien Chiang / Yao-Kai Chang / Wei-Han Hui / Shu-Wei Chang / Chen-Yi Liao / Yi-Chuan Chang / Chun-Jung Chen / Wei-Chen Wang / Chien-Chen Lai / Chun-Hsiung Wang / Siou-Ying Luo / Ya- ...Authors: Wesley Tien Chiang / Yao-Kai Chang / Wei-Han Hui / Shu-Wei Chang / Chen-Yi Liao / Yi-Chuan Chang / Chun-Jung Chen / Wei-Chen Wang / Chien-Chen Lai / Chun-Hsiung Wang / Siou-Ying Luo / Ya-Ping Huang / Shan-Ho Chou / Tzyy-Leng Horng / Ming-Hon Hou / Stephen P Muench / Ren-Shiang Chen / Ming-Daw Tsai / Nien-Jen Hu /
Abstract: The K uptake system KtrAB is essential for bacterial survival in low K environments. The activity of KtrAB is regulated by nucleotides and Na. Previous studies proposed a putative gating mechanism of ...The K uptake system KtrAB is essential for bacterial survival in low K environments. The activity of KtrAB is regulated by nucleotides and Na. Previous studies proposed a putative gating mechanism of KtrB regulated by KtrA upon binding to ATP or ADP. However, how Na activates KtrAB and the Na binding site remain unknown. Here we present the cryo-EM structures of ATP- and ADP-bound KtrAB from Bacillus subtilis (BsKtrAB) both solved at 2.8 Å. A cryo-EM density at the intra-dimer interface of ATP-KtrA was identified as Na, as supported by X-ray crystallography and ICP-MS. Thermostability assays and functional studies demonstrated that Na binding stabilizes the ATP-bound BsKtrAB complex and enhances its K flux activity. Comparing ATP- and ADP-BsKtrAB structures suggests that BsKtrB Arg417 and Phe91 serve as a channel gate. The synergism of ATP and Na in activating BsKtrAB is likely applicable to Na-activated K channels in central nervous system.
History
DepositionJul 10, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Apr 3, 2024Provider: repository / Type: Initial release
Revision 1.1May 15, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.title / _citation.year
Revision 1.2May 22, 2024Group: Database references / Category: citation / citation_author
Item: _citation.page_first / _citation.page_last ..._citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Ktr system potassium uptake protein A
B: Ktr system potassium uptake protein A
hetero molecules


Theoretical massNumber of molelcules
Total (without water)51,0525
Polymers49,8342
Non-polymers1,2193
Water00
1
A: Ktr system potassium uptake protein A
B: Ktr system potassium uptake protein A
hetero molecules

A: Ktr system potassium uptake protein A
B: Ktr system potassium uptake protein A
hetero molecules

A: Ktr system potassium uptake protein A
B: Ktr system potassium uptake protein A
hetero molecules

A: Ktr system potassium uptake protein A
B: Ktr system potassium uptake protein A
hetero molecules


Theoretical massNumber of molelcules
Total (without water)204,20920
Polymers199,3348
Non-polymers4,87512
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_555-x,-y,z1
crystal symmetry operation3_555-y,x,z1
crystal symmetry operation4_555y,-x,z1
Buried area30470 Å2
ΔGint-199 kcal/mol
Surface area73250 Å2
MethodPISA
Unit cell
Length a, b, c (Å)122.418, 122.418, 83.853
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number79
Space group name H-MI4
Space group name HallI4
Symmetry operation#1: x,y,z
#2: -y,x,z
#3: y,-x,z
#4: -x,-y,z
#5: x+1/2,y+1/2,z+1/2
#6: -y+1/2,x+1/2,z+1/2
#7: y+1/2,-x+1/2,z+1/2
#8: -x+1/2,-y+1/2,z+1/2

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Components

#1: Protein Ktr system potassium uptake protein A / K(+)-uptake protein KtrA


Mass: 24916.760 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus subtilis (bacteria) / Gene: ktrA, yuaA, BSU31090 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: O32080
#2: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: ATP, energy-carrying molecule*YM
#3: Chemical ChemComp-TL / THALLIUM (I) ION


Mass: 204.383 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Tl / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 3.15 Å3/Da / Density % sol: 60.98 %
Crystal growTemperature: 293.15 K / Method: vapor diffusion, hanging drop / pH: 7.5
Details: 100 mM HEPES-KOH pH 7.5, 3% PEG 6000, 2.5% 2-methyl-2,4-pentanediol, 5 mM Thallium(I) acetate

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Data collection

DiffractionMean temperature: 80 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: NSRRC / Beamline: TPS 05A / Wavelength: 1 Å
DetectorType: RAYONIX MX300-HS / Detector: CCD / Date: May 22, 2022
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 3.1→24.01 Å / Num. obs: 22017 / % possible obs: 99.71 % / Redundancy: 15.2 % / Biso Wilson estimate: 101.12 Å2 / Rpim(I) all: 0.01 / Rrim(I) all: 0.04 / Net I/σ(I): 55.87
Reflection shellResolution: 3.1→3.16 Å / Mean I/σ(I) obs: 5.57 / Num. unique obs: 1134 / CC1/2: 0.97 / CC star: 0.992 / Rpim(I) all: 0.123

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Processing

Software
NameVersionClassification
PHENIX1.20.1_4487refinement
Blu-Icedata collection
HKL-2000data reduction
HKL-2000data scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 3.1→24.01 Å / SU ML: 0.4629 / Cross valid method: FREE R-VALUE / σ(F): 1.35 / Phase error: 33.5757
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflection
Rfree0.2618 2202 10 %
Rwork0.2367 19815 -
obs0.2394 22017 99.71 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.1 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 110.86 Å2
Refinement stepCycle: LAST / Resolution: 3.1→24.01 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3412 0 63 0 3475
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00233530
X-RAY DIFFRACTIONf_angle_d0.49794782
X-RAY DIFFRACTIONf_chiral_restr0.0429552
X-RAY DIFFRACTIONf_plane_restr0.0031600
X-RAY DIFFRACTIONf_dihedral_angle_d6.2976470
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.1-3.160.39821280.41451210X-RAY DIFFRACTION100
3.17-3.240.33571370.3791277X-RAY DIFFRACTION99.72
3.24-3.320.40731400.3591254X-RAY DIFFRACTION100
3.32-3.410.36581380.31811213X-RAY DIFFRACTION99.93
3.41-3.510.34971360.32351239X-RAY DIFFRACTION100
3.51-3.620.37431460.3251231X-RAY DIFFRACTION99.93
3.62-3.750.38411360.30071226X-RAY DIFFRACTION100
3.75-3.90.31931430.29671250X-RAY DIFFRACTION99.93
3.9-4.080.27011420.27651232X-RAY DIFFRACTION99.78
4.08-4.290.24451390.25371236X-RAY DIFFRACTION99.93
4.29-4.560.26361430.21651258X-RAY DIFFRACTION100
4.56-4.90.22541380.21511240X-RAY DIFFRACTION99.86
4.91-5.390.2751330.23041247X-RAY DIFFRACTION100
5.4-6.160.2481380.22641244X-RAY DIFFRACTION100
6.17-7.710.27781350.21061244X-RAY DIFFRACTION99.86
7.73-24.010.14261300.14251214X-RAY DIFFRACTION96.48
Refinement TLS params.Method: refined / Origin x: -17.972308756 Å / Origin y: 33.4867164416 Å / Origin z: -34.0711319934 Å
111213212223313233
T0.668987332498 Å20.120918284198 Å2-0.00820058944965 Å2-0.801012676107 Å2-0.260753080995 Å2--0.714148090921 Å2
L2.46502273891 °2-0.660245801688 °2-0.312342605499 °2-2.91729268184 °2-1.02298009898 °2--2.48351126595 °2
S-0.553818972883 Å °-0.0562733695236 Å °0.175296942152 Å °0.232116665023 Å °0.289399672451 Å °-0.387121278212 Å °-0.242886524756 Å °-0.424696437395 Å °0.268068987017 Å °
Refinement TLS groupSelection details: all

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