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- PDB-8h1o: Cryo-EM structure of KpFtsZ-monobody double helical tube -

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Basic information

Entry
Database: PDB / ID: 8h1o
TitleCryo-EM structure of KpFtsZ-monobody double helical tube
Components
  • Cell division protein FtsZ
  • Mb(Ec/KpFtsZ_S1)
KeywordsCELL CYCLE / bacterial cell division / divisome / FtsZ / monobody / tubulin
Function / homology
Function and homology information


FtsZ-dependent cytokinesis / division septum assembly / cell division site / protein polymerization / GTPase activity / GTP binding / cytoplasm
Similarity search - Function
Tubulin-like protein FtsZ/CetZ / Cell division protein FtsZ / Cell division protein FtsZ, conserved site / Cell division protein FtsZ, C-terminal / FtsZ family, C-terminal domain / FtsZ protein signature 1. / FtsZ protein signature 2. / Tubulin/FtsZ family, C-terminal domain / Tubulin/FtsZ-like, C-terminal domain / Tubulin/FtsZ, C-terminal ...Tubulin-like protein FtsZ/CetZ / Cell division protein FtsZ / Cell division protein FtsZ, conserved site / Cell division protein FtsZ, C-terminal / FtsZ family, C-terminal domain / FtsZ protein signature 1. / FtsZ protein signature 2. / Tubulin/FtsZ family, C-terminal domain / Tubulin/FtsZ-like, C-terminal domain / Tubulin/FtsZ, C-terminal / Tubulin/FtsZ, 2-layer sandwich domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ, GTPase domain / Tubulin/FtsZ, GTPase domain superfamily
Similarity search - Domain/homology
GUANOSINE-5'-DIPHOSPHATE / Cell division protein FtsZ
Similarity search - Component
Biological speciesKlebsiella pneumoniae (bacteria)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 2.67 Å
AuthorsFujita, J. / Amesaka, H. / Yoshizawa, T. / Kuroda, N. / Kamimura, N. / Hara, M. / Inoue, T. / Namba, K. / Tanaka, S. / Matsumura, H.
Funding support Japan, 16items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)JP18K05445 Japan
Japan Society for the Promotion of Science (JSPS)JP21K05386 Japan
Japan Society for the Promotion of Science (JSPS)JP18K06094 Japan
Japan Society for the Promotion of Science (JSPS)JP19H04735 Japan
Japan Society for the Promotion of Science (JSPS)JP20K22630 Japan
the Japan Science Society2018-3011 Japan
the Japan Science Society2022-4052 Japan
Japan Science and TechnologyJPMJOP1861 Japan
Japan Agency for Medical Research and Development (AMED)JP21am0101117 Japan
Japan Agency for Medical Research and Development (AMED)JP22ama121003 Japan
Japan Agency for Medical Research and Development (AMED)JP17pc0101020 Japan
JEOL YOKOGUSHI Research Alliance Laboratories of Osaka University Japan
Institute for Protein Research, Osaka UniversityCR-20-02 Japan
Institute for Protein Research, Osaka UniversityCR-21-02 Japan
Japan Agency for Medical Research and Development (AMED)JP21am0101070 Japan
G-7 Scholarship Foundation
CitationJournal: Nat Commun / Year: 2023
Title: Structures of a FtsZ single protofilament and a double-helical tube in complex with a monobody.
Authors: Junso Fujita / Hiroshi Amesaka / Takuya Yoshizawa / Kota Hibino / Natsuki Kamimura / Natsuko Kuroda / Takamoto Konishi / Yuki Kato / Mizuho Hara / Tsuyoshi Inoue / Keiichi Namba / Shun-Ichi ...Authors: Junso Fujita / Hiroshi Amesaka / Takuya Yoshizawa / Kota Hibino / Natsuki Kamimura / Natsuko Kuroda / Takamoto Konishi / Yuki Kato / Mizuho Hara / Tsuyoshi Inoue / Keiichi Namba / Shun-Ichi Tanaka / Hiroyoshi Matsumura /
Abstract: FtsZ polymerizes into protofilaments to form the Z-ring that acts as a scaffold for accessory proteins during cell division. Structures of FtsZ have been previously solved, but detailed mechanistic ...FtsZ polymerizes into protofilaments to form the Z-ring that acts as a scaffold for accessory proteins during cell division. Structures of FtsZ have been previously solved, but detailed mechanistic insights are lacking. Here, we determine the cryoEM structure of a single protofilament of FtsZ from Klebsiella pneumoniae (KpFtsZ) in a polymerization-preferred conformation. We also develop a monobody (Mb) that binds to KpFtsZ and FtsZ from Escherichia coli without affecting their GTPase activity. Crystal structures of the FtsZ-Mb complexes reveal the Mb binding mode, while addition of Mb in vivo inhibits cell division. A cryoEM structure of a double-helical tube of KpFtsZ-Mb at 2.7 Å resolution shows two parallel protofilaments. Our present study highlights the physiological roles of the conformational changes of FtsZ in treadmilling that regulate cell division.
History
DepositionOct 3, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Aug 2, 2023Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Cell division protein FtsZ
B: Mb(Ec/KpFtsZ_S1)
hetero molecules


Theoretical massNumber of molelcules
Total (without water)50,8003
Polymers50,3572
Non-polymers4431
Water0
1
A: Cell division protein FtsZ
B: Mb(Ec/KpFtsZ_S1)
hetero molecules
x 100


Theoretical massNumber of molelcules
Total (without water)5,080,000300
Polymers5,035,680200
Non-polymers44,320100
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
helical symmetry operation99

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Components

#1: Protein Cell division protein FtsZ /


Mass: 40574.926 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Klebsiella pneumoniae (bacteria) / Production host: Escherichia coli (E. coli) / References: UniProt: W9BCK7
#2: Protein Mb(Ec/KpFtsZ_S1)


Mass: 9781.873 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)
#3: Chemical ChemComp-GDP / GUANOSINE-5'-DIPHOSPHATE / Guanosine diphosphate


Type: RNA linking / Mass: 443.201 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H15N5O11P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: GDP, energy-carrying molecule*YM
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: FILAMENT / 3D reconstruction method: helical reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1Cryo-EM structure of KpFtsZ-monobody double helical tubeCOMPLEX#1-#20MULTIPLE SOURCES
2KpFtsZCOMPLEX#11RECOMBINANT
3monobodyCOMPLEX#21RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Klebsiella pneumoniae (bacteria)573
33Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
22Escherichia coli (E. coli)562
33Escherichia coli (E. coli)562
Buffer solutionpH: 7.5
Details: 1 mM GMPPNP and 0.12 mM PC190723 were supplemented.
Buffer component
IDConc.NameFormulaBuffer-ID
120 mM2-[4-(2-Hydroxyethyl)-1-piperazinyl]ethanesulfonic acidHEPES1
2100 mMpotassium chlorideKCl1
325 mMsodium chlorideNaClSodium chloride1
45 mMmagnesium chlorideMgCl21
SpecimenConc.: 4 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: 1.2x molar excess of Mb was supplemented.
Specimen supportDetails: The graphene grid was chemically oxidized and modified.
Grid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE

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Electron microscopy imaging

MicroscopyModel: JEOL CRYO ARM 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 60000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: JEOL CRYOSPECPORTER
Image recordingAverage exposure time: 3 sec. / Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1
EM imaging opticsEnergyfilter name: In-column Omega Filter / Energyfilter slit width: 20 eV

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
EM software
IDNameVersionCategory
1cryoSPARC3.3.1particle selection
2SerialEM3.8image acquisition
4cryoSPARC3.3.1CTF correction
7UCSF Chimera1.15model fitting
9cryoSPARC3.3.1initial Euler assignment
10cryoSPARC3.3.1final Euler assignment
12cryoSPARC3.3.13D reconstruction
13PHENIX1.19.2model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Helical symmertyAngular rotation/subunit: -23.398 ° / Axial rise/subunit: 7.703 Å / Axial symmetry: C2
Particle selectionNum. of particles selected: 181247
3D reconstructionResolution: 2.67 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 90711 / Algorithm: FOURIER SPACE / Symmetry type: HELICAL
Atomic model buildingSpace: REAL
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 34.49 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00333015
ELECTRON MICROSCOPYf_angle_d0.5264099
ELECTRON MICROSCOPYf_chiral_restr0.0451490
ELECTRON MICROSCOPYf_plane_restr0.0031534
ELECTRON MICROSCOPYf_dihedral_angle_d7.3154442

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