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- PDB-9s35: Cryo-EM structure of Candida albicans Vrg4 bound to an inhibitory... -

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Basic information

Entry
Database: PDB / ID: 9s35
TitleCryo-EM structure of Candida albicans Vrg4 bound to an inhibitory nanobody.
Components
  • GDP-mannose transporter
  • Llama Nanobody
KeywordsTRANSPORT PROTEIN / GDP-Mannose transporter / Membrane Protein / Nucleotide Sugar Transporter
Function / homology
Function and homology information


GDP-mannose transmembrane transporter activity / GDP-mannose transmembrane transport / hyphal growth / protein N-linked glycosylation via asparagine / antiporter activity / cytoplasmic vesicle membrane / Golgi membrane / endoplasmic reticulum membrane / Golgi apparatus
Similarity search - Function
GDP-mannose transporter
Similarity search - Component
Biological speciesLama glama (llama)
Candida albicans (yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å
AuthorsDeme, J.C. / Parker, J.L. / Lea, S.M. / Newstead, S.
Funding support United States, United Kingdom, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)Intramural Research Program United States
Wellcome Trust215519/Z/19/Z United Kingdom
Medical Research Council (MRC, United Kingdom)MR/S021043/1 United Kingdom
CitationJournal: Res Sq / Year: 2025
Title: Structural basis for transport and inhibition of nucleotide sugar transport in pathogenic fungi.
Authors: Joanne L Parker / Justin C Deme / Bjarne Feddersen / Susan M Lea / Simon Newstead /
Abstract: GDP-Mannose transporters are Golgi-localised solute carriers that are essential for the virulence of pathogenic fungi, serving as critical components of fungal glycosylation pathways. However, the ...GDP-Mannose transporters are Golgi-localised solute carriers that are essential for the virulence of pathogenic fungi, serving as critical components of fungal glycosylation pathways. However, the mechanism by which nucleotide sugars are recognised and transported across the Golgi membrane remains unclear, hindering efforts to develop effective inhibitors that could serve as novel antifungal agents. Here, we present cryo-EM structures of the GDP-Mannose transporter, Vrg4, from in complex with nanobodies and in both the cytoplasmic and Golgi-facing states. Structural comparisons between these two states, in addition to a GDP-mannose bound structure, demonstrate the importance of ligand movement during transport. Additionally, we demonstrate the ability of the nanobodies to specifically inhibit Vrg4, presenting proof-of-principle that nanobodies can be used as effective inhibitors of nucleotide sugar transport and glycosylation in cells.
History
DepositionJul 23, 2025Deposition site: PDBE / Processing site: PDBE
Revision 1.0Sep 3, 2025Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: Llama Nanobody
A: GDP-mannose transporter


Theoretical massNumber of molelcules
Total (without water)50,5592
Polymers50,5592
Non-polymers00
Water1267
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Antibody Llama Nanobody


Mass: 13376.000 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lama glama (llama) / Production host: Escherichia coli BL21(DE3) (bacteria)
#2: Protein GDP-mannose transporter / GMT


Mass: 37183.223 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Candida albicans (yeast) / Gene: VRG4, CAALFM_C107700CA, CaO19.1232, CaO19.8817 / Production host: Saccharomyces cerevisiae (brewer's yeast) / Strain (production host): BJ5460 / References: UniProt: Q5A477
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 7 / Source method: isolated from a natural source / Formula: H2O
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Nanobody bound to C.albicans Vrg4 transporter / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Candida albicans (yeast)
Source (recombinant)Organism: Saccharomyces cerevisiae (brewer's yeast)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 55 e/Å2 / Film or detector model: TFS FALCON 4i (4k x 4k)

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Processing

EM software
IDNameVersionCategory
1SIMPLEparticle selection
2PHENIX1.21.2_5419model refinement
13cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 348612 / Symmetry type: POINT
RefinementHighest resolution: 3 Å
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0043434
ELECTRON MICROSCOPYf_angle_d0.4884668
ELECTRON MICROSCOPYf_dihedral_angle_d3.989471
ELECTRON MICROSCOPYf_chiral_restr0.039541
ELECTRON MICROSCOPYf_plane_restr0.006570

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