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- EMDB-19981: Cryo-EM structure of the full-length Pseudomonas aeruginosa bacte... -

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Basic information

Entry
Database: EMDB / ID: EMD-19981
TitleCryo-EM structure of the full-length Pseudomonas aeruginosa bacteriophytochrome in its Pr state
Map dataComposite map generated with phenix combine maps.
Sample
  • Complex: Homodimeric complex of the protein bacteriophytochrome containing its cofactor biliverdin
    • Protein or peptide: Bacteriophytochrome
  • Ligand: 3-[2-[(Z)-[3-(2-carboxyethyl)-5-[(Z)-(4-ethenyl-3-methyl-5-oxidanylidene-pyrrol-2-ylidene)methyl]-4-methyl-pyrrol-1-ium -2-ylidene]methyl]-5-[(Z)-[(3E)-3-ethylidene-4-methyl-5-oxidanylidene-pyrrolidin-2-ylidene]methyl]-4-methyl-1H-pyrrol-3- yl]propanoic acid
KeywordsPhotosensor / Photoreceptor / Phytochrome / Bacterial protein / CYTOSOLIC PROTEIN
Function / homology
Function and homology information


osmosensory signaling via phosphorelay pathway / detection of visible light / phosphorelay response regulator activity / protein kinase activator activity / histidine kinase / phosphorelay sensor kinase activity / photoreceptor activity / regulation of DNA-templated transcription / ATP binding / identical protein binding
Similarity search - Function
: / Phytochrome / Phytochrome, PHY domain superfamily / PAS fold-2 / PAS fold / Phytochrome, central region / Phytochrome region / Phytochrome chromophore attachment domain / Phytochrome chromophore attachment site domain profile. / His Kinase A (phospho-acceptor) domain ...: / Phytochrome / Phytochrome, PHY domain superfamily / PAS fold-2 / PAS fold / Phytochrome, central region / Phytochrome region / Phytochrome chromophore attachment domain / Phytochrome chromophore attachment site domain profile. / His Kinase A (phospho-acceptor) domain / His Kinase A (phosphoacceptor) domain / Signal transduction histidine kinase, dimerisation/phosphoacceptor domain / Signal transduction histidine kinase, dimerisation/phosphoacceptor domain superfamily / Histidine kinase domain / Histidine kinase domain profile. / GAF domain / Domain present in phytochromes and cGMP-specific phosphodiesterases. / GAF domain / GAF-like domain superfamily / Histidine kinase-, DNA gyrase B-, and HSP90-like ATPase / Histidine kinase-like ATPases / PAS domain superfamily / Histidine kinase/HSP90-like ATPase / Histidine kinase/HSP90-like ATPase superfamily
Similarity search - Domain/homology
Biological speciesPseudomonas aeruginosa (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.95 Å
AuthorsBodizs S / Westenhoff S
Funding support Sweden, 1 items
OrganizationGrant numberCountry
Swedish Research Council Sweden
CitationJournal: Structure / Year: 2024
Title: Cryo-EM structures of a bathy phytochrome histidine kinase reveal a unique light-dependent activation mechanism.
Authors: Szabolcs Bódizs / Petra Mészáros / Lukas Grunewald / Heikki Takala / Sebastian Westenhoff /
Abstract: Phytochromes are photoreceptor proteins in plants, fungi, and bacteria. They can adopt two photochromic states with differential biochemical responses. The structural changes transducing the signal ...Phytochromes are photoreceptor proteins in plants, fungi, and bacteria. They can adopt two photochromic states with differential biochemical responses. The structural changes transducing the signal from the chromophore to the biochemical output modules are poorly understood due to challenges in capturing structures of the dynamic, full-length protein. Here, we present cryoelectron microscopy (cryo-EM) structures of the phytochrome from Pseudomonas aeruginosa (PaBphP) in its resting (Pfr) and photoactivated (Pr) state. The kinase-active Pr state has an asymmetric, dimeric structure, whereas the kinase-inactive Pfr state opens up. This behavior is different from other known phytochromes and we explain it with the unusually short connection between the photosensory and output modules. Multiple sequence alignment of this region suggests evolutionary optimization for different modes of signal transduction in sensor proteins. The results establish a new mechanism for light-sensing by phytochrome histidine kinases and provide input for the design of optogenetic phytochrome variants.
History
DepositionMar 28, 2024-
Header (metadata) releaseSep 4, 2024-
Map releaseSep 4, 2024-
UpdateSep 11, 2024-
Current statusSep 11, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19981.png

Downloads & links

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Map

FileDownload / File: emd_19981.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationComposite map generated with phenix combine maps.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 400 pix.
= 331.2 Å		0.83 Å/pix.
x 400 pix.
= 331.2 Å		0.83 Å/pix.
x 400 pix.
= 331.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.828 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 10.0
Minimum - Maximum-56.351813999999997 - 83.353874000000005
Average (Standard dev.)0.015432564 (±1.0491204)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 331.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Homodimeric complex of the protein bacteriophytochrome containing...

EntireName: Homodimeric complex of the protein bacteriophytochrome containing its cofactor biliverdin
Components
  • Complex: Homodimeric complex of the protein bacteriophytochrome containing its cofactor biliverdin
    • Protein or peptide: Bacteriophytochrome
  • Ligand: 3-[2-[(Z)-[3-(2-carboxyethyl)-5-[(Z)-(4-ethenyl-3-methyl-5-oxidanylidene-pyrrol-2-ylidene)methyl]-4-methyl-pyrrol-1-ium -2-ylidene]methyl]-5-[(Z)-[(3E)-3-ethylidene-4-methyl-5-oxidanylidene-pyrrolidin-2-ylidene]methyl]-4-methyl-1H-pyrrol-3- yl]propanoic acid

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Supramolecule #1: Homodimeric complex of the protein bacteriophytochrome containing...

SupramoleculeName: Homodimeric complex of the protein bacteriophytochrome containing its cofactor biliverdin
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Pseudomonas aeruginosa (bacteria) / Location in cell: cytoplasm
Molecular weightTheoretical: 160 KDa

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Macromolecule #1: Bacteriophytochrome

MacromoleculeName: Bacteriophytochrome / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: histidine kinase
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 82.094867 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MTSITPVTLA NCEDEPIHVP GAIQPHGALV TLRADGMVLA ASENIQALLG FVASPGSYLT QEQVGPEVLR MLEEGLTGNG PWSNSVETR IGEHLFDVIG HSYKEVFYLE FEIRTADTLS ITSFTLNAQR IIAQVQLHND TASLLSNVTD ELRRMTGYDR V MAYRFRHD ...String:
MTSITPVTLA NCEDEPIHVP GAIQPHGALV TLRADGMVLA ASENIQALLG FVASPGSYLT QEQVGPEVLR MLEEGLTGNG PWSNSVETR IGEHLFDVIG HSYKEVFYLE FEIRTADTLS ITSFTLNAQR IIAQVQLHND TASLLSNVTD ELRRMTGYDR V MAYRFRHD DSGEVVAESR REDLESYLGQ RYPASDIPAQ ARRLYIQNPI RLIADVAYTP MRVFPALNPE TNESFDLSYS VL RSVSPIH CEYLTNMGVR ASMSISIVVG GKLWGLFSCH HMSPKLIPYP VRMSFQIFSQ VCSAIVERLE QGRIAELLRV STE RRLALA RRARDADDLF GALAHPDDGI AALIPCDGAL VMLGGRTLSI RGDFERQAGN VLQRLQRDPE RDIYHTDNWP QPSE DSPDG GDCCGVLAIR FHRQESGWIF WFRHEEVHRI RWGGKPEKLL TIGPSGPRLT PRGSFEAWEE VVRGHSTPWS ETDLA IAEK LRLDLMELCL NHAAEVDRMR QRLIAVLGHD LRNPLQSISM AAALLSSSDT RTTELRQHIS ASSSRMERLV SQILDM SRL QSGIGLTVNP VDTDVSQLVR QIVCETDVAY PGLVIEIAID PQVRAVVDPD RYAQVAANLL SNARHHGLPG RPVLVTL TR QGDEVCLSVL NETSGLSEAQ LANLFEPFKR ESADNQRNRN GLGIGLYISQ AIAQAHQGRI DVDCRDDVIT FCLRLPVR Q AETGSSSLEH HHHHH

UniProtKB: Bacteriophytochrome

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Macromolecule #2: 3-[2-[(Z)-[3-(2-carboxyethyl)-5-[(Z)-(4-ethenyl-3-methyl-5-oxidan...

MacromoleculeName: 3-[2-[(Z)-[3-(2-carboxyethyl)-5-[(Z)-(4-ethenyl-3-methyl-5-oxidanylidene-pyrrol-2-ylidene)methyl]-4-methyl-pyrrol-1-ium -2-ylidene]methyl]-5-[(Z)-[(3E)-3-ethylidene-4-methyl-5-oxidanylidene- ...Name: 3-[2-[(Z)-[3-(2-carboxyethyl)-5-[(Z)-(4-ethenyl-3-methyl-5-oxidanylidene-pyrrol-2-ylidene)methyl]-4-methyl-pyrrol-1-ium -2-ylidene]methyl]-5-[(Z)-[(3E)-3-ethylidene-4-methyl-5-oxidanylidene-pyrrolidin-2-ylidene]methyl]-4-methyl-1H-pyrrol-3- yl]propanoic acid
type: ligand / ID: 2 / Number of copies: 2 / Formula: LBV
Molecular weightTheoretical: 585.67 Da

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1.5 mg/mL
BufferpH: 7.8
Component:
ConcentrationFormulaName
80.0 mMC4H11NO3Tris
10.0 mMMgCl2Magnesium chloride
150.0 mMCH3CO2KPotassium acetate

Details: 80 mM Tris, 10 mM MgCl2, 150 mM CH3CO2K, pH 7.8
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Support film - Film thickness: 50 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: OTHER / Pretreatment - Pressure: 0.04 kPa / Details: Current: 20 mA
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: Blotting for 5 s from both sides.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 2 / Number real images: 30336 / Average exposure time: 1.7 sec. / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 2.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.95 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. v4.2.1)
Details: Final map is a composite, input map 1 has a reported resolution of 2.95 A, map 2 has a reported resolution of 4.31 A
Number images used: 889461
Initial angle assignmentType: NOT APPLICABLE / Software - Name: cryoSPARC (ver. v4.2.1)
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
Output model

PDB-9eut:
Cryo-EM structure of the full-length Pseudomonas aeruginosa bacteriophytochrome in its Pr state

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