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- PDB-7r1v: Crystal structure of E.coli BamA beta-barrel in complex with dyno... -

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Basic information

Entry
Database: PDB / ID: 7r1v
TitleCrystal structure of E.coli BamA beta-barrel in complex with dynobactin A
Components
  • Dynobactin A
  • Outer membrane protein assembly factor BamA
KeywordsMEMBRANE PROTEIN / Beta-Barrel / outer membrane / protein insertion / protein folding / protein maturation / antibiotic / natural product / cyclized peptide
Function / homology
Function and homology information


Bam protein complex / Gram-negative-bacterium-type cell outer membrane assembly / protein insertion into membrane
Similarity search - Function
Outer membrane protein assembly factor BamA / POTRA domain, BamA/TamA-like / Surface antigen variable number repeat / POTRA domain / POTRA domain profile. / Surface antigen D15-like / Bacterial surface antigen (D15) / Omp85 superfamily domain
Similarity search - Domain/homology
: / Uncharacterized protein / Outer membrane protein assembly factor BamA
Similarity search - Component
Biological speciesEscherichia coli O157:H7 (bacteria)
Photorhabdus australis (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.5 Å
AuthorsJakob, R.P. / Hiller, S. / Maier, T.
Funding support Switzerland, 1items
OrganizationGrant numberCountry
Swiss National Science Foundation177084 Switzerland
CitationJournal: Nat Microbiol / Year: 2022
Title: Computational identification of a systemic antibiotic for gram-negative bacteria.
Authors: Ryan D Miller / Akira Iinishi / Seyed Majed Modaresi / Byung-Kuk Yoo / Thomas D Curtis / Patrick J Lariviere / Libang Liang / Sangkeun Son / Samantha Nicolau / Rachel Bargabos / Madeleine ...Authors: Ryan D Miller / Akira Iinishi / Seyed Majed Modaresi / Byung-Kuk Yoo / Thomas D Curtis / Patrick J Lariviere / Libang Liang / Sangkeun Son / Samantha Nicolau / Rachel Bargabos / Madeleine Morrissette / Michael F Gates / Norman Pitt / Roman P Jakob / Parthasarathi Rath / Timm Maier / Andrey G Malyutin / Jens T Kaiser / Samantha Niles / Blake Karavas / Meghan Ghiglieri / Sarah E J Bowman / Douglas C Rees / Sebastian Hiller / Kim Lewis /
Abstract: Discovery of antibiotics acting against Gram-negative species is uniquely challenging due to their restrictive penetration barrier. BamA, which inserts proteins into the outer membrane, is an ...Discovery of antibiotics acting against Gram-negative species is uniquely challenging due to their restrictive penetration barrier. BamA, which inserts proteins into the outer membrane, is an attractive target due to its surface location. Darobactins produced by Photorhabdus, a nematode gut microbiome symbiont, target BamA. We reasoned that a computational search for genes only distantly related to the darobactin operon may lead to novel compounds. Following this clue, we identified dynobactin A, a novel peptide antibiotic from Photorhabdus australis containing two unlinked rings. Dynobactin is structurally unrelated to darobactins, but also targets BamA. Based on a BamA-dynobactin co-crystal structure and a BAM-complex-dynobactin cryo-EM structure, we show that dynobactin binds to the BamA lateral gate, uniquely protruding into its β-barrel lumen. Dynobactin showed efficacy in a mouse systemic Escherichia coli infection. This study demonstrates the utility of computational approaches to antibiotic discovery and suggests that dynobactin is a promising lead for drug development.
History
DepositionFeb 3, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Sep 28, 2022Provider: repository / Type: Initial release
Revision 1.1Nov 30, 2022Group: 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.pdbx_database_id_DOI / _citation.title / _citation.year
Revision 1.2Dec 7, 2022Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.3Jan 31, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model
Revision 1.4Apr 3, 2024Group: Database references / Derived calculations ...Database references / Derived calculations / Source and taxonomy / Structure summary
Category: pdbx_entity_src_syn / pdbx_entry_details ...pdbx_entity_src_syn / pdbx_entry_details / pdbx_struct_assembly / pdbx_struct_assembly_prop / struct_ref / struct_ref_seq
Item: _pdbx_entity_src_syn.ncbi_taxonomy_id / _pdbx_entity_src_syn.organism_scientific ..._pdbx_entity_src_syn.ncbi_taxonomy_id / _pdbx_entity_src_syn.organism_scientific / _pdbx_entry_details.compound_details / _pdbx_struct_assembly.details / _pdbx_struct_assembly.method_details / _struct_ref.db_code / _struct_ref.db_name / _struct_ref.pdbx_align_begin / _struct_ref.pdbx_db_accession / _struct_ref.pdbx_seq_one_letter_code / _struct_ref_seq.db_align_beg / _struct_ref_seq.db_align_end / _struct_ref_seq.pdbx_db_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Outer membrane protein assembly factor BamA
B: Dynobactin A


Theoretical massNumber of molelcules
Total (without water)45,1172
Polymers45,1172
Non-polymers00
Water1,38777
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area1130 Å2
ΔGint-5 kcal/mol
Surface area18310 Å2
MethodPISA
Unit cell
Length a, b, c (Å)65.790, 71.324, 116.555
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121

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Components

#1: Protein Outer membrane protein assembly factor BamA


Mass: 43805.457 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli O157:H7 (bacteria) / Gene: bamA, yaeT, Z0188, ECs0179 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P0A942
#2: Protein/peptide Dynobactin A


Type: Peptide-like / Class: Antibiotic / Mass: 1311.405 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Photorhabdus australis (bacteria) / References: UniProt: A0A1C0U7H2, BIRD: PRD_002404
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 77 / Source method: isolated from a natural source / Formula: H2O
Compound detailsDynobactin A is a decapeptide (1-WNSNVHSYRF-10) with 2 closed rings. One carbon-carbon bond formed ...Dynobactin A is a decapeptide (1-WNSNVHSYRF-10) with 2 closed rings. One carbon-carbon bond formed between the C6 of Trp (W1) and the beta-carbon of Asn (N4). Second an unusual nitrogen-carbon linkage formed between the imidazole N-Epsilon-2 of His (H6) and the beta-carbon of Tyr (Y8).
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: 3.12 Å3/Da / Density % sol: 60.59 %
Crystal growTemperature: 298 K / Method: vapor diffusion
Details: 0.12 M lithium sulfate, 0.02 M Tris pH7.5, 0.1 M sodium citrate pH 5.0, and 20% (v/v) PEG300

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SLS / Beamline: X06DA / Wavelength: 1.000003 Å
DetectorType: DECTRIS PILATUS 2M-F / Detector: PIXEL / Date: Dec 18, 2020
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.000003 Å / Relative weight: 1
ReflectionResolution: 2.5→48.36 Å / Num. obs: 19622 / % possible obs: 99 % / Redundancy: 27.9 % / CC1/2: 0.998 / Rmerge(I) obs: 0.217 / Rpim(I) all: 0.044 / Net I/σ(I): 9
Reflection shellResolution: 2.5→2.6 Å / Rmerge(I) obs: 4.571 / Mean I/σ(I) obs: 1 / Num. unique obs: 2171 / CC1/2: 0.718 / Rpim(I) all: 0.893

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Processing

Software
NameVersionClassification
BUSTER2.10.3 (18-SEP-2020)refinement
PDB_EXTRACT3.27data extraction
XDSdata reduction
Aimlessdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 7NRF

7nrf


Resolution: 2.5→48.36 Å / Cor.coef. Fo:Fc: 0.884 / Cor.coef. Fo:Fc free: 0.906 / Cross valid method: THROUGHOUT / σ(F): 0 / SU R Blow DPI: 0.406 / SU Rfree Blow DPI: 0.27
Details: HYDROGENS WERE FULLY REFINED WITH ZERO OCCUPANCY AT NUCLEAR POSITION. REFINEMENT NOTES. NUMBER OF REFINEMENT NOTES : 1 NOTE 1 : IDEAL-DIST CONTACT TERM CONTACT SETUP. ALL ATOMS HAVE CCP4 ATOM TYPE FROM LIBRARY
RfactorNum. reflection% reflectionSelection details
Rfree0.2748 985 5.04 %RANDOM
Rwork0.2465 ---
obs0.248 19555 99.7 %-
Displacement parametersBiso max: 310.76 Å2 / Biso mean: 113.32 Å2 / Biso min: 41.78 Å2
Baniso -1Baniso -2Baniso -3
1-24.2839 Å20 Å20 Å2
2--4.9408 Å20 Å2
3----29.2247 Å2
Refine analyzeLuzzati coordinate error obs: 0.51 Å
Refinement stepCycle: final / Resolution: 2.5→48.36 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2986 0 0 77 3063
Biso mean---68.9 -
Num. residues----377
Refine LS restraints
Refine-IDTypeNumberRestraint functionWeightDev ideal
X-RAY DIFFRACTIONt_dihedral_angle_d1661SINUSOIDAL2
X-RAY DIFFRACTIONt_trig_c_planes
X-RAY DIFFRACTIONt_gen_planes953HARMONIC5
X-RAY DIFFRACTIONt_it3080HARMONIC10
X-RAY DIFFRACTIONt_nbd
X-RAY DIFFRACTIONt_improper_torsion
X-RAY DIFFRACTIONt_pseud_angle
X-RAY DIFFRACTIONt_chiral_improper_torsion376SEMIHARMONIC5
X-RAY DIFFRACTIONt_sum_occupancies
X-RAY DIFFRACTIONt_utility_distance
X-RAY DIFFRACTIONt_utility_angle
X-RAY DIFFRACTIONt_utility_torsion
X-RAY DIFFRACTIONt_ideal_dist_contact3953SEMIHARMONIC4
X-RAY DIFFRACTIONt_bond_d5787HARMONIC20.008
X-RAY DIFFRACTIONt_angle_deg10294HARMONIC20.93
X-RAY DIFFRACTIONt_omega_torsion3.19
X-RAY DIFFRACTIONt_other_torsion16.1
LS refinement shellResolution: 2.5→2.52 Å / Rfactor Rfree error: 0 / Total num. of bins used: 50
RfactorNum. reflection% reflection
Rfree0.4144 20 5 %
Rwork0.2505 380 -
all0.2602 400 -
obs--99.49 %
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
17.7279-0.6473-1.87444.67910.115712.24490.0717-1.0520.24890.73020.14950.2809-0.83230.5849-0.2212-0.3931-0.06180.1026-0.3598-0.01290.77183.11098.365140.5182
23.07270.13430.07434.3127-1.71270.5068-0.1861-0.6095-0.39970.8144-0.0417-0.25710.01620.07550.2278-0.4993-0.0109-0.0852-0.357-0.01920.737312.311418.257237.6747
33.80570.0118-0.36852.8219-0.40030.50.08270.2052-0.044-0.0907-0.0362-0.28060.0207-0.0498-0.0465-0.42720.02070.0285-0.35490.00610.59158.899325.240420.6522
413.03715.3211-6.21315.7125-3.85957.5492-0.16641.1215-0.5252-0.520.48220.42520.3704-0.6516-0.3158-0.48480.0678-0.0539-0.425-0.1320.5508-4.34117.218614.7744
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1{ A|423 - A|454 }A423 - 454
2X-RAY DIFFRACTION2{ A|455 - A|537 }A455 - 537
3X-RAY DIFFRACTION3{ A|538 - A|766 }A538 - 766
4X-RAY DIFFRACTION4{ A|767 - A|809 }A767 - 809

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