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- EMDB-36882: Cryo-EM structure of nucleotide-bound ComA with ZinC ion -

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Basic information

Entry
Database: EMDB / ID: EMD-36882
TitleCryo-EM structure of nucleotide-bound ComA with ZinC ion
Map data
Sample
  • Complex: ComA
    • Protein or peptide: Competence factor transporting ATP-binding protein/permease ComA
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: ZINC ION
KeywordsABC transport / PCAT / MEMBRANE PROTEIN / TRANSPORT PROTEIN
Function / homology
Function and homology information


ABC-type bacteriocin transporter activity / cysteine-type peptidase activity / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / ATP hydrolysis activity / proteolysis / ATP binding / membrane
Similarity search - Function
Peptidase C39, ABC-type bacteriocin transporter / Peptidase C39 family / Peptidase C39, bacteriocin processing / Peptidase family C39 domain profile. / Type 1 protein exporter / ABC transporter transmembrane region / ABC transporter type 1, transmembrane domain / ABC transporter integral membrane type-1 fused domain profile. / ABC transporter type 1, transmembrane domain superfamily / ABC transporter-like, conserved site ...Peptidase C39, ABC-type bacteriocin transporter / Peptidase C39 family / Peptidase C39, bacteriocin processing / Peptidase family C39 domain profile. / Type 1 protein exporter / ABC transporter transmembrane region / ABC transporter type 1, transmembrane domain / ABC transporter integral membrane type-1 fused domain profile. / ABC transporter type 1, transmembrane domain superfamily / ABC transporter-like, conserved site / ABC transporters family signature. / ABC transporter / ABC transporter-like, ATP-binding domain / ATP-binding cassette, ABC transporter-type domain profile. / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Competence factor transporting ATP-binding protein/permease ComA
Similarity search - Component
Biological speciesStreptococcus pneumoniae (bacteria) / Streptococcus pneumoniae D39 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.9 Å
AuthorsYu L / Xin X / Min L / Feng H
Funding support Singapore, 1 items
OrganizationGrant numberCountry
Ministry of Education (MoE, Singapore)A-0008412-00-00 Singapore
CitationJournal: Nat Commun / Year: 2023
Title: Structural basis of peptide secretion for Quorum sensing by ComA.
Authors: Lin Yu / Xin Xu / Wan-Zhen Chua / Hao Feng / Zheng Ser / Kai Shao / Jian Shi / Yumei Wang / Zongli Li / Radoslaw M Sobota / Lok-To Sham / Min Luo /
Abstract: Quorum sensing (QS) is a crucial regulatory mechanism controlling bacterial signalling and holds promise for novel therapies against antimicrobial resistance. In Gram-positive bacteria, such as ...Quorum sensing (QS) is a crucial regulatory mechanism controlling bacterial signalling and holds promise for novel therapies against antimicrobial resistance. In Gram-positive bacteria, such as Streptococcus pneumoniae, ComA is a conserved efflux pump responsible for the maturation and secretion of peptide signals, including the competence-stimulating peptide (CSP), yet its structure and function remain unclear. Here, we functionally characterize ComA as an ABC transporter with high ATP affinity and determined its cryo-EM structures in the presence or absence of CSP or nucleotides. Our findings reveal a network of strong electrostatic interactions unique to ComA at the intracellular gate, a putative binding pocket for two CSP molecules, and negatively charged residues facilitating CSP translocation. Mutations of these residues affect ComA's peptidase activity in-vitro and prevent CSP export in-vivo. We demonstrate that ATP-Mg triggers the outward-facing conformation of ComA for CSP release, rather than ATP alone. Our study provides molecular insights into the QS signal peptide secretion, highlighting potential targets for QS-targeting drugs.
History
DepositionJul 17, 2023-
Header (metadata) releaseOct 11, 2023-
Map releaseOct 11, 2023-
UpdateMay 8, 2024-
Current statusMay 8, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_36882.png

Downloads & links

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Map

FileDownload / File: emd_36882.map.gz / Format: CCP4 / Size: 42.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.11 Å/pix.
x 224 pix.
= 247.52 Å		1.11 Å/pix.
x 224 pix.
= 247.52 Å		1.11 Å/pix.
x 224 pix.
= 247.52 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.105 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.2
Minimum - Maximum-0.5880383 - 0.8266945
Average (Standard dev.)0.00042129823 (±0.02856815)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions224224224
Spacing224224224
CellA=B=C: 247.52 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_36882_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_36882_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : ComA

EntireName: ComA
Components
  • Complex: ComA
    • Protein or peptide: Competence factor transporting ATP-binding protein/permease ComA
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: ZINC ION

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Supramolecule #1: ComA

SupramoleculeName: ComA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Streptococcus pneumoniae (bacteria)

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Macromolecule #1: Competence factor transporting ATP-binding protein/permease ComA

MacromoleculeName: Competence factor transporting ATP-binding protein/permease ComA
type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus pneumoniae D39 (bacteria) / Strain: D39
Molecular weightTheoretical: 80.433031 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MKFGKRHYRP QVDQMDCGVA SLAMVFGYYG SYYFLAHLRE LAKTTMDGTT ALGLVKVAEE IGFETRAIKA DMTLFDLPDL TFPFVAHVL KEGKLLHYYV VTGQDKDSIH IADPDPGVKL TKLPRERFEE EWTGVTLFMA PSPDYKPHKE QKNGLLSFIP I LVKQRGLI ...String:
MKFGKRHYRP QVDQMDCGVA SLAMVFGYYG SYYFLAHLRE LAKTTMDGTT ALGLVKVAEE IGFETRAIKA DMTLFDLPDL TFPFVAHVL KEGKLLHYYV VTGQDKDSIH IADPDPGVKL TKLPRERFEE EWTGVTLFMA PSPDYKPHKE QKNGLLSFIP I LVKQRGLI ANIVLATLLV TVINIVGSYY LQSIIDTYVP DQMRSTLGII SIGLVIVYIL QQILSYAQEY LLLVLGQRLS ID VILSYIK HVFHLPMSFF ATRRTGEIVS RFTDANSIID ALASTILSIF LDVSTVVIIS LVLFSQNTNL FFMTLLALPI YTV IIFAFM KPFEKMNRDT MEANAVLSSS IIEDINGIET IKSLTSESQR YQKIDKEFVD YLKKSFTYSR AESQQKALKK VAHL LLNVG ILWMGAVLVM DGKMSLGQLI TYNTLLVYFT NPLENIINLQ TKLQTAQVAN NRLNEVYLVA SEFEEKKTVE DLSLM KGDM TFKQVHYKYG YGRDVLSDIN LTVPQGSKVA FVGISGSGKT TLAKMMVNFY DPSQGEISLG GVNLNQIDKK ALRQYI NYL PQQPYVFNGT ILENLLLGAK EGTTQEDILR AVELAEIRED IERMPLNYQT ELTSDGAGIS GGQRQRIALA RALLTDA PV IILDEATSSL DILTEKRIVD NLIALDKTLI FIAHRLTIAE RTEKVVVLDQ GKIVEEGKHA DLLAQGGFYA HLVNS

UniProtKB: Competence factor transporting ATP-binding protein/permease ComA

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Macromolecule #2: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 2 / Number of copies: 2 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

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Macromolecule #3: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 3 / Number of copies: 2 / Formula: ZN
Molecular weightTheoretical: 65.409 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

Concentration5 mg/mL
BufferpH: 7.5 / Details: 25 mM Tris, pH 7.5, 150 mM NaCl, 2 mM DTT
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 1.2 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 50384
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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