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- PDB-6y4l: Crystal structure of human ER membrane protein complex subunits E... -

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Basic information

Entry
Database: PDB / ID: 6y4l
TitleCrystal structure of human ER membrane protein complex subunits EMC2 and EMC9
Components(ER membrane protein complex subunit ...Endoplasmic reticulum) x 2
KeywordsPROTEIN TRANSPORT / Complex / insertase
Function / homology
Function and homology information


extrinsic component of endoplasmic reticulum membrane / EMC complex / protein insertion into ER membrane by stop-transfer membrane-anchor sequence / tail-anchored membrane protein insertion into ER membrane / endoplasmic reticulum membrane / endoplasmic reticulum / cytoplasm
Similarity search - Function
ER membrane protein complex subunit 8/9 / Uncharacterised protein family (UPF0172) / ER membrane protein complex subunit 2-like / Tetratricopeptide repeat / TPR repeat region circular profile. / TPR repeat profile. / MPN domain / MPN domain profile. / Tetratricopeptide repeats / Tetratricopeptide repeat / Tetratricopeptide-like helical domain superfamily
Similarity search - Domain/homology
DI(HYDROXYETHYL)ETHER / ER membrane protein complex subunit 2 / ER membrane protein complex subunit 9
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / SAD / Resolution: 2.2 Å
AuthorsHegde, R.S. / O'Donnell, J.P.
Funding support United Kingdom, Germany, 2items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MC_UP_A022_1007 United Kingdom
European Molecular Biology Organization (EMBO)ALTF-18-2018 Germany
CitationJournal: Elife / Year: 2020
Title: The architecture of EMC reveals a path for membrane protein insertion.
Authors: John P O'Donnell / Ben P Phillips / Yuichi Yagita / Szymon Juszkiewicz / Armin Wagner / Duccio Malinverni / Robert J Keenan / Elizabeth A Miller / Ramanujan S Hegde /
Abstract: Approximately 25% of eukaryotic genes code for integral membrane proteins that are assembled at the endoplasmic reticulum. An abundant and widely conserved multi-protein complex termed EMC has been ...Approximately 25% of eukaryotic genes code for integral membrane proteins that are assembled at the endoplasmic reticulum. An abundant and widely conserved multi-protein complex termed EMC has been implicated in membrane protein biogenesis, but its mechanism of action is poorly understood. Here, we define the composition and architecture of human EMC using biochemical assays, crystallography of individual subunits, site-specific photocrosslinking, and cryo-EM reconstruction. Our results suggest that EMC's cytosolic domain contains a large, moderately hydrophobic vestibule that can bind a substrate's transmembrane domain (TMD). The cytosolic vestibule leads into a lumenally-sealed, lipid-exposed intramembrane groove large enough to accommodate a single substrate TMD. A gap between the cytosolic vestibule and intramembrane groove provides a potential path for substrate egress from EMC. These findings suggest how EMC facilitates energy-independent membrane insertion of TMDs, explain why only short lumenal domains are translocated by EMC, and constrain models of EMC's proposed chaperone function.
History
DepositionFeb 21, 2020Deposition site: PDBE / Processing site: PDBE
Revision 1.0May 27, 2020Provider: repository / Type: Initial release
Revision 1.1Jun 10, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_abbrev / _citation.journal_id_ISSN ..._citation.journal_abbrev / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: ER membrane protein complex subunit 2
B: ER membrane protein complex subunit 9
hetero molecules


Theoretical massNumber of molelcules
Total (without water)57,28112
Polymers56,1342
Non-polymers1,14710
Water1,44180
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: light scattering
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area5280 Å2
ΔGint-32 kcal/mol
Surface area20790 Å2
MethodPISA
Unit cell
Length a, b, c (Å)52.577, 84.939, 122.585
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121
Space group name HallP2ac2ab
Symmetry operation#1: x,y,z
#2: x+1/2,-y+1/2,-z
#3: -x,y+1/2,-z+1/2
#4: -x+1/2,-y,z+1/2

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Components

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ER membrane protein complex subunit ... , 2 types, 2 molecules AB

#1: Protein ER membrane protein complex subunit 2 / Endoplasmic reticulum / Tetratricopeptide repeat protein 35 / TPR repeat protein 35


Mass: 32352.562 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC2, KIAA0103, TTC35 / Production host: Escherichia coli (E. coli) / References: UniProt: Q15006
#2: Protein ER membrane protein complex subunit 9 / Endoplasmic reticulum / Protein FAM158A


Mass: 23781.264 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC9, C14orf122, FAM158A, CGI-112 / Production host: Escherichia coli (E. coli) / References: UniProt: Q9Y3B6

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Non-polymers , 5 types, 90 molecules

#3: Chemical ChemComp-SO4 / SULFATE ION / Sulfate


Mass: 96.063 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: SO4
#4: Chemical ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL / Ethylene glycol


Mass: 62.068 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C2H6O2
#5: Chemical ChemComp-PE4 / 2-{2-[2-(2-{2-[2-(2-ETHOXY-ETHOXY)-ETHOXY]-ETHOXY}-ETHOXY)-ETHOXY]-ETHOXY}-ETHANOL / POLYETHYLENE GLYCOL PEG4000 / Polyethylene glycol


Mass: 354.436 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Formula: C16H34O8 / Comment: precipitant*YM
#6: Chemical ChemComp-PEG / DI(HYDROXYETHYL)ETHER / Diethylene glycol


Mass: 106.120 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C4H10O3
#7: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 80 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 2

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Sample preparation

Crystal
IDDensity Matthews (Å3/Da)Density % sol (%)
12.4449.67
2
Crystal grow
Temperature (K)Crystal-IDMethodpHDetails
293.151vapor diffusion, hanging drop8.334% PEG4000, 0.1 M Tris pH 8.3, and 0.2 M lithium sulfate (For I03 native dataset)
293.152vapor diffusion, hanging drop8.328% PEG4000, 0.1 M Tris pH 8.3, and 0.2 M lithium sulfate (For I23 S-SAD dataset)

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Data collection

Diffraction
IDMean temperature (K)Crystal-IDSerial crystal experiment
11001N
21
Diffraction source
SourceSiteBeamlineIDWavelength (Å)
SYNCHROTRONDiamond I0310.9763
SYNCHROTRONDiamond I2322.755
Detector
TypeIDDetectorDate
DECTRIS EIGER2 X 16M1PIXELJul 13, 2019
DECTRIS PILATUS 12M2PIXELJul 3, 2019
Radiation
IDProtocolMonochromatic (M) / Laue (L)Scattering typeWavelength-ID
1SINGLE WAVELENGTHMx-ray1
2SINGLE WAVELENGTHMx-ray2
Radiation wavelength
IDWavelength (Å)Relative weight
10.97631
22.7551
Reflection

Entry-ID: 6Y4L

Resolution (Å)Num. obs% possible obs (%)Redundancy (%)CC1/2Diffraction-IDNet I/σ(I)
2.2-49.72866810051.91117.6
2.65-49.61646899.740.61236.8
Reflection shell
Resolution (Å)Mean I/σ(I) obsNum. unique obsCC1/2Diffraction-ID
2.2-2.271.924330.9261
2.65-2.781.821090.8172

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Processing

Software
NameVersionClassification
PHENIX1.17.1_3660refinement
DIALSdata reduction
PHENIXphasing
Aimlessdata scaling
RefinementMethod to determine structure: SAD
Starting model: XXXX

Resolution: 2.2→49.7 Å / Cross valid method: FREE R-VALUE
RfactorNum. reflection% reflection
Rfree0.2499 1995 6.99 %
Rwork0.2031 --
obs0.2064 28558 99.81 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso mean: 70.84 Å2
Refinement stepCycle: LAST / Resolution: 2.2→49.7 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3446 0 72 80 3598
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00723592
X-RAY DIFFRACTIONf_angle_d0.84374877
X-RAY DIFFRACTIONf_chiral_restr0.0484541
X-RAY DIFFRACTIONf_plane_restr0.0053628
X-RAY DIFFRACTIONf_dihedral_angle_d11.6878519
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)
17.367433500720.3740136471561.57331940735.79383275967-1.611119831496.10949568708-0.2794450788360.4087732877090.669515233548-0.235903073639-0.309246575084-0.547280144146-0.8975069462181.010482768380.5195858436580.617132924659-0.0103125989015-0.01838710339210.6232649495330.07617448005710.42339858737724.9338007206-19.5219189637-22.5740742037
22.74310024563-0.879976342136-0.6187317830673.400141892820.3379852310263.698201400040.0533678184396-0.2348817229090.02217258608560.191753550046-0.1832094482370.0227743669053-0.08420363082810.1729589240560.1299640084080.298206275824-0.0276336071716-0.01325880604790.4343282270340.03248564479350.359632077201-0.115409302406-7.020917862072.7333362223
32.999938956360.243721736293-0.1561035454024.74334945454-0.6324940926914.508813250930.2170142094720.2996214841850.318325073625-0.629407530672-0.194964676787-0.237880383983-0.6594139306620.114408274602-0.05879327360470.5710544665550.05678812882840.08177429665960.3931856468690.04083955698550.469683964123-0.2275461676287.42364368362-16.1093550962
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection details
1X-RAY DIFFRACTION1(chain A and resid 11:97)
2X-RAY DIFFRACTION2(chain A and resid 98:278)
3X-RAY DIFFRACTION3(chain B and resid 2:194)

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