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- PDB-8em4: Cryo-EM structure of LRP2 at pH 7.5 -

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Basic information

Entry
Database: PDB / ID: 8em4
TitleCryo-EM structure of LRP2 at pH 7.5
ComponentsLow-density lipoprotein receptor-related protein 2
KeywordsMEMBRANE PROTEIN / LRP2 / Megalin / GP330 / Endocytosis
Function / homology
Function and homology information


Transport of RCbl within the body / endocytic hemoglobin import into cell / diol metabolic process / chemoattraction of axon / positive regulation of lipoprotein transport / pulmonary artery morphogenesis / secondary heart field specification / Retinoid metabolism and transport / positive regulation of oligodendrocyte progenitor proliferation / folate import across plasma membrane ...Transport of RCbl within the body / endocytic hemoglobin import into cell / diol metabolic process / chemoattraction of axon / positive regulation of lipoprotein transport / pulmonary artery morphogenesis / secondary heart field specification / Retinoid metabolism and transport / positive regulation of oligodendrocyte progenitor proliferation / folate import across plasma membrane / metal ion transport / response to leptin / ventricular compact myocardium morphogenesis / protein transporter activity / Cargo recognition for clathrin-mediated endocytosis / hormone binding / vitamin D metabolic process / neuron projection arborization / Clathrin-mediated endocytosis / hormone secretion / coronary artery morphogenesis / outflow tract septum morphogenesis / coronary vasculature development / insulin-like growth factor I binding / transcytosis / protein import / aorta development / cargo receptor activity / ventricular septum development / positive regulation of neurogenesis / endosomal transport / low-density lipoprotein particle receptor binding / hemoglobin binding / amyloid-beta clearance / positive regulation of endocytosis / brush border / vagina development / negative regulation of BMP signaling pathway / response to X-ray / endocytic vesicle / axonal growth cone / clathrin-coated pit / forebrain development / receptor-mediated endocytosis / kidney development / neural tube closure / PDZ domain binding / endosome lumen / nuclear receptor binding / brush border membrane / sensory perception of sound / cellular response to growth factor stimulus / SH3 domain binding / endocytosis / male gonad development / protein transport / apical part of cell / heart development / protein-folding chaperone binding / cell population proliferation / positive regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / receptor complex / endosome / membrane raft / apical plasma membrane / external side of plasma membrane / axon / calcium ion binding / dendrite / protein-containing complex binding / negative regulation of apoptotic process / Golgi apparatus / cell surface / endoplasmic reticulum / protein-containing complex / extracellular space / membrane / plasma membrane / cytoplasm
Similarity search - Function
Complement Clr-like EGF domain / Complement Clr-like EGF-like / Low-density lipoprotein receptor repeat class B / LDL-receptor class B (LDLRB) repeat profile. / LDLR class B repeat / Low-density lipoprotein-receptor YWTD domain / Low-density lipoprotein receptor domain class A / Low-density lipoprotein (LDL) receptor class A, conserved site / LDL-receptor class A (LDLRA) domain signature. / LDL-receptor class A (LDLRA) domain profile. ...Complement Clr-like EGF domain / Complement Clr-like EGF-like / Low-density lipoprotein receptor repeat class B / LDL-receptor class B (LDLRB) repeat profile. / LDLR class B repeat / Low-density lipoprotein-receptor YWTD domain / Low-density lipoprotein receptor domain class A / Low-density lipoprotein (LDL) receptor class A, conserved site / LDL-receptor class A (LDLRA) domain signature. / LDL-receptor class A (LDLRA) domain profile. / Calcium-binding EGF domain / Low-density lipoprotein receptor domain class A / Low-density lipoprotein (LDL) receptor class A repeat / LDL receptor-like superfamily / Six-bladed beta-propeller, TolB-like / EGF-type aspartate/asparagine hydroxylation site / EGF-like calcium-binding, conserved site / Calcium-binding EGF-like domain signature. / Aspartic acid and asparagine hydroxylation site. / EGF-like calcium-binding domain / Calcium-binding EGF-like domain / Epidermal growth factor-like domain. / EGF-like domain profile. / Growth factor receptor cysteine-rich domain superfamily / EGF-like domain signature 2. / EGF-like domain signature 1. / EGF-like domain
Similarity search - Domain/homology
2-acetamido-2-deoxy-beta-D-galactopyranose / Low-density lipoprotein receptor-related protein 2
Similarity search - Component
Biological speciesMus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.83 Å
AuthorsBeenken, A. / Cerutti, G. / Brasch, J. / Fitzpatrick, A.W. / Barasch, J. / Shapiro, L.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)DK124667 United States
CitationJournal: Cell / Year: 2023
Title: Structures of LRP2 reveal a molecular machine for endocytosis.
Authors: Andrew Beenken / Gabriele Cerutti / Julia Brasch / Yicheng Guo / Zizhang Sheng / Hediye Erdjument-Bromage / Zainab Aziz / Shelief Y Robbins-Juarez / Estefania Y Chavez / Goran Ahlsen / ...Authors: Andrew Beenken / Gabriele Cerutti / Julia Brasch / Yicheng Guo / Zizhang Sheng / Hediye Erdjument-Bromage / Zainab Aziz / Shelief Y Robbins-Juarez / Estefania Y Chavez / Goran Ahlsen / Phinikoula S Katsamba / Thomas A Neubert / Anthony W P Fitzpatrick / Jonathan Barasch / Lawrence Shapiro /
Abstract: The low-density lipoprotein (LDL) receptor-related protein 2 (LRP2 or megalin) is representative of the phylogenetically conserved subfamily of giant LDL receptor-related proteins, which function in ...The low-density lipoprotein (LDL) receptor-related protein 2 (LRP2 or megalin) is representative of the phylogenetically conserved subfamily of giant LDL receptor-related proteins, which function in endocytosis and are implicated in diseases of the kidney and brain. Here, we report high-resolution cryoelectron microscopy structures of LRP2 isolated from mouse kidney, at extracellular and endosomal pH. The structures reveal LRP2 to be a molecular machine that adopts a conformation for ligand binding at the cell surface and for ligand shedding in the endosome. LRP2 forms a homodimer, the conformational transformation of which is governed by pH-sensitive sites at both homodimer and intra-protomer interfaces. A subset of LRP2 deleterious missense variants in humans appears to impair homodimer assembly. These observations lay the foundation for further understanding the function and mechanism of LDL receptors and implicate homodimerization as a conserved feature of the LRP receptor subfamily.
History
DepositionSep 26, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Feb 8, 2023Provider: repository / Type: Initial release
Revision 1.1Feb 22, 2023Group: Database references / Structure summary / Category: audit_author / citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Mar 8, 2023Group: Database references / Category: citation / Item: _citation.journal_volume / _citation.page_first

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Low-density lipoprotein receptor-related protein 2
B: Low-density lipoprotein receptor-related protein 2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,063,617178
Polymers1,039,4932
Non-polymers24,124176
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, equilibrium centrifugation, light scattering
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Low-density lipoprotein receptor-related protein 2 / LRP-2 / Glycoprotein 330 / gp330 / Megalin


Mass: 519746.500 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: A2ARV4
#2: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 2 / Source method: isolated from a natural source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}}LINUCSPDB-CARE
#3: Sugar...
ChemComp-NGA / 2-acetamido-2-deoxy-beta-D-galactopyranose / N-acetyl-beta-D-galactosamine / 2-acetamido-2-deoxy-beta-D-galactose / 2-acetamido-2-deoxy-D-galactose / 2-acetamido-2-deoxy-galactose / N-ACETYL-D-GALACTOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 32 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
IdentifierTypeProgram
DGalpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-galactopyranosamineCOMMON NAMEGMML 1.0
b-D-GalpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GalNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
#4: Sugar...
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 58 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
#5: Chemical...
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 84 / Source method: obtained synthetically / Formula: Ca / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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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: LRP2 at neutral pH / Type: COMPLEX / Details: Endogenously purified from mouse kidney / Entity ID: #1 / Source: NATURAL
Molecular weightExperimental value: NO
Source (natural)Organism: Mus musculus (house mouse)
Buffer solutionpH: 7.5
SpecimenConc.: 2 mg/ml / Embedding 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: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1500 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 54.87 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.17.1_3660refinement
PHENIX1.17.1_3660refinement
ISOLDE1.2.2model building
UCSF ChimeraX1.2/v9model building
EM software
IDNameCategory
2Leginonimage acquisition
4cryoSPARCCTF correction
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
13cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 2.83 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 492737 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT

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