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- PDB-7v55: Crystal structure of phospholipase D from Pseudomonas aeruginosa ... -

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Basic information

Entry
Database: PDB / ID: 7v55
TitleCrystal structure of phospholipase D from Pseudomonas aeruginosa PAO1 using in situ proteolysis
ComponentsPhospholipase D
KeywordsHYDROLASE / phospholipase D / Toxin
Function / homologyPhospholipase D family / Phospholipase D Active site motif / Phospholipase D. Active site motifs. / Phospholipase D/Transphosphatidylase / Phospholipase D phosphodiesterase active site profile. / phospholipase D activity / phospholipid catabolic process / Phospholipase D
Function and homology information
Biological speciesPseudomonas aeruginosa (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3 Å
AuthorsYang, Y. / Li, Z.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Nat Commun / Year: 2022
Title: Structural insights into PA3488-mediated inactivation of Pseudomonas aeruginosa PldA.
Authors: Xiaoyun Yang / Zongqiang Li / Liang Zhao / Zhun She / Zengqiang Gao / Sen-Fang Sui / Yuhui Dong / Yanhua Li /
Abstract: PldA, a phospholipase D (PLD) effector, catalyzes hydrolysis of the phosphodiester bonds of glycerophospholipids-the main component of cell membranes-and assists the invasion of the opportunistic ...PldA, a phospholipase D (PLD) effector, catalyzes hydrolysis of the phosphodiester bonds of glycerophospholipids-the main component of cell membranes-and assists the invasion of the opportunistic pathogen Pseudomonas aeruginosa. As a cognate immunity protein, PA3488 can inhibit the activity of PldA to avoid self-toxicity. However, the precise inhibitory mechanism remains elusive. We determine the crystal structures of full-length and truncated PldA and the cryogenic electron microscopy structure of the PldA-PA3488 complex. Structural analysis reveals that there are different intermediates of PldA between the "open" and "closed" states of the catalytic pocket, accompanied by significant conformational changes in the "lid" region and the peripheral helical domain. Through structure-based mutational analysis, we identify the key residues responsible for the enzymatic activity of PldA. Together, these data provide an insight into the molecular mechanisms of PldA invasion and its neutralization by PA3488, aiding future design of PLD-targeted inhibitors and drugs.
History
DepositionAug 16, 2021Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Aug 17, 2022Provider: repository / Type: Initial release
Revision 1.1Mar 1, 2023Group: 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.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Nov 29, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Phospholipase D
hetero molecules


Theoretical massNumber of molelcules
Total (without water)122,5182
Polymers122,4781
Non-polymers401
Water543
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)83.566, 83.566, 242.335
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number152
Space group name H-MP3121

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Components

#1: Protein Phospholipase D /


Mass: 122478.062 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Gene: pldA, PA3487 / Production host: Escherichia coli (E. coli) / References: UniProt: Q9HYC2
#2: Chemical ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Ca
#3: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestN

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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: 1.99 Å3/Da / Density % sol: 38.33 %
Crystal growTemperature: 291.1 K / Method: evaporation
Details: 0.2M Potassium phosphate dibasic, 20% w/v Polyethylene glycol 3350

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SSRF / Beamline: BL17U1 / Wavelength: 0.97853 Å
DetectorType: ADSC QUANTUM 315r / Detector: CCD / Date: Apr 26, 2019
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97853 Å / Relative weight: 1
ReflectionResolution: 3→72.37 Å / Num. obs: 19242 / % possible obs: 94.4 % / Redundancy: 5.3 % / Biso Wilson estimate: 68.27 Å2 / CC1/2: 0.996 / Rmerge(I) obs: 0.123 / Rpim(I) all: 0.057 / Rrim(I) all: 0.136 / Net I/σ(I): 11.8 / Num. measured all: 102295 / Scaling rejects: 85
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsNum. measured allNum. unique obsCC1/2Rpim(I) allRrim(I) allNet I/σ(I) obs% possible all
3-3.185.60.7341782432050.7460.3360.8112.299.3
9-72.374.60.02438488310.9990.0120.02742.897.5

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Processing

Software
NameVersionClassification
Aimless0.7.4data scaling
PHENIX1.19_4092refinement
PDB_EXTRACT3.27data extraction
XDSdata reduction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 7V53

7v53


Resolution: 3→69.34 Å / SU ML: 0.35 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 24.47 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.249 929 4.83 %
Rwork0.1893 18297 -
obs0.1923 19226 93.99 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 152.93 Å2 / Biso mean: 60.7261 Å2 / Biso min: 25.26 Å2
Refinement stepCycle: final / Resolution: 3→69.34 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6118 0 1 3 6122
Biso mean--72.99 47.28 -
Num. residues----781
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 7

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
3-3.160.3381470.26622672281999
3.16-3.360.28521210.236627522873100
3.36-3.620.32191340.22942731286599
3.62-3.980.2521960.20471774187065
3.98-4.550.21461340.16982731286599
4.56-5.740.23591490.16322759290899
5.74-69.340.21561480.16772878302697

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