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- PDB-8g2v: Cryo-EM structure of recombinant human LECT2 amyloid fibril core -

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Basic information

Entry
Database: PDB / ID: 8g2v
TitleCryo-EM structure of recombinant human LECT2 amyloid fibril core
ComponentsLeukocyte cell-derived chemotaxin-2
KeywordsPROTEIN FIBRIL / amyloid / LECT2 / human / recombinant / fibril / protein / ALECT2 / cryo-EM
Function / homology
Function and homology information


skeletal system development / chemotaxis / extracellular space / identical protein binding / metal ion binding / cytoplasm
Similarity search - Function
Leukocyte cell-derived chemotaxin 2 / Leukocyte cell-derived chemotaxin 2, chordata / Peptidase M23 / Peptidase family M23 / Duplicated hybrid motif
Similarity search - Domain/homology
Leukocyte cell-derived chemotaxin-2
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 2.715 Å
AuthorsRichards, L.S. / Flores, M.D. / Zink, S. / Schibrowsky, N.A. / Sawaya, M.R. / Rodriguez, J.A.
Funding support United States, 5items
OrganizationGrant numberCountry
Department of Energy (DOE, United States)DE-FC02-02ER63421 United States
National Science Foundation (NSF, United States)DMR-1548924 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)Grant R35 GM128867 United States
National Institutes of Health/National Center for Research Resources (NIH/NCRR)5T32GM008496 United States
National Institutes of Health/National Center for Research Resources (NIH/NCRR)U24 GM129541 United States
CitationJournal: Structure / Year: 2023
Title: Cryo-EM structure of a human LECT2 amyloid fibril reveals a network of polar ladders at its core.
Authors: Logan S Richards / Maria D Flores / Samantha Zink / Natalie A Schibrowsky / Michael R Sawaya / Jose A Rodriguez /
Abstract: ALECT2 systemic amyloidosis is associated with deposition of the leukocyte cell-derived chemotaxin-2 (LECT2) protein in the form of fibrils. In ALECT2 amyloidosis, ALECT2 fibrils deposit in the ...ALECT2 systemic amyloidosis is associated with deposition of the leukocyte cell-derived chemotaxin-2 (LECT2) protein in the form of fibrils. In ALECT2 amyloidosis, ALECT2 fibrils deposit in the glomerulus, resulting in renal failure. Patients lack effective treatment options outside of renal transplant or dialysis. The structure of globular LECT2 has been determined but structures of ALECT2 amyloid fibrils remain unknown. Using single-particle cryo-EM, we find that recombinant human LECT2 forms robust twisting fibrils with canonical amyloid features. ALECT2 fibrils contain two mating protofilaments spanning residues 55-75 of the LECT2 sequence. The geometry of the ALECT2 fibril displays features in line with other pathogenic amyloids. Its core is tightly packed and stabilized by both hydrophobic contacts and hydrogen-bonded uncharged polar residues. The robustness of ALECT2 fibril cores is illustrated by their resistance to denaturants and proteases. This ALECT2 fibril structure presents a potential new target for treatments against ALECT2 systemic amyloidosis.
History
DepositionFeb 6, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 16, 2023Provider: repository / Type: Initial release
Revision 1.1Sep 13, 2023Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed ..._citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Nov 15, 2023Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Leukocyte cell-derived chemotaxin-2
B: Leukocyte cell-derived chemotaxin-2
C: Leukocyte cell-derived chemotaxin-2
D: Leukocyte cell-derived chemotaxin-2
E: Leukocyte cell-derived chemotaxin-2
F: Leukocyte cell-derived chemotaxin-2
G: Leukocyte cell-derived chemotaxin-2
H: Leukocyte cell-derived chemotaxin-2
I: Leukocyte cell-derived chemotaxin-2
J: Leukocyte cell-derived chemotaxin-2


Theoretical massNumber of molelcules
Total (without water)23,89810
Polymers23,89810
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: mass spectrometry, native gel electrophoresis
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein/peptide
Leukocyte cell-derived chemotaxin-2 / LECT-2 / hLECT2


Mass: 2389.752 Da / Num. of mol.: 10
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: LECT2 / Production host: Escherichia coli (E. coli) / References: UniProt: O14960

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: FILAMENT / 3D reconstruction method: helical reconstruction

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Sample preparation

ComponentName: Recombinant human LECT2 amyloid fibril core. / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 6.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1800 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 1.3 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)
EM imaging opticsPhase plate: VOLTA PHASE PLATE

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Processing

SoftwareName: PHENIX / Version: 1.20_4459: / Classification: refinement
EM software
IDNameVersionCategory
1crYOLOparticle selection
2EPUimage acquisition
4CTFFIND4CTF correction
7Cootmodel fitting
10RELION3.1final Euler assignment
11RELION3.1classification
12RELION3.13D reconstruction
13PHENIXmodel refinement
CTF correctionType: NONE
Helical symmertyAngular rotation/subunit: 179.49 ° / Axial rise/subunit: 2.345 Å / Axial symmetry: C1
Particle selectionNum. of particles selected: 1622942
3D reconstructionResolution: 2.715 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 24770 / Symmetry type: HELICAL
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL

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