+Open data
-Basic information
Entry | Database: PDB / ID: 6x58 | ||||||
---|---|---|---|---|---|---|---|
Title | MPER-Fluc-Ec2 bound to 10E8v4 antibody | ||||||
Components |
| ||||||
Keywords | MEMBRANE PROTEIN / antibody / chaperone / MEMBRANE PROTEIN-IMMUNE SYSTEM complex | ||||||
Function / homology | Function and homology information fluoride channel activity / cellular detoxification of fluoride / positive regulation of plasma membrane raft polarization / positive regulation of receptor clustering / positive regulation of establishment of T cell polarity / host cell endosome membrane / clathrin-dependent endocytosis of virus by host cell / viral protein processing / fusion of virus membrane with host plasma membrane / virus-mediated perturbation of host defense response ...fluoride channel activity / cellular detoxification of fluoride / positive regulation of plasma membrane raft polarization / positive regulation of receptor clustering / positive regulation of establishment of T cell polarity / host cell endosome membrane / clathrin-dependent endocytosis of virus by host cell / viral protein processing / fusion of virus membrane with host plasma membrane / virus-mediated perturbation of host defense response / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / host cell plasma membrane / structural molecule activity / virion membrane / membrane / metal ion binding / plasma membrane Similarity search - Function | ||||||
Biological species | Homo sapiens (human) Human immunodeficiency virus Escherichia coli (E. coli) | ||||||
Method | X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.26 Å | ||||||
Authors | McIlwain, B.C. / Stockbridge, R.B. | ||||||
Funding support | United States, 1items
| ||||||
Citation | Journal: J Mol Biol / Year: 2021 Title: N-terminal Transmembrane-Helix Epitope Tag for X-ray Crystallography and Electron Microscopy of Small Membrane Proteins. Authors: Benjamin C McIlwain / Amanda L Erwin / Alexander R Davis / B Ben Koff / Louise Chang / Tatsiana Bylund / Gwo-Yu Chuang / Peter D Kwong / Melanie D Ohi / Yen-Ting Lai / Randy B Stockbridge / Abstract: Structural studies of membrane proteins, especially small membrane proteins, are associated with well-known experimental challenges. Complexation with monoclonal antibody fragments is a common ...Structural studies of membrane proteins, especially small membrane proteins, are associated with well-known experimental challenges. Complexation with monoclonal antibody fragments is a common strategy to augment such proteins; however, generating antibody fragments that specifically bind a target protein is not trivial. Here we identify a helical epitope, from the membrane-proximal external region (MPER) of the gp41-transmembrane subunit of the HIV envelope protein, that is recognized by several well-characterized antibodies and that can be fused as a contiguous extension of the N-terminal transmembrane helix of a broad range of membrane proteins. To analyze whether this MPER-epitope tag might aid structural studies of small membrane proteins, we determined an X-ray crystal structure of a membrane protein target that does not crystallize without the aid of crystallization chaperones, the Fluc fluoride channel, fused to the MPER epitope and in complex with antibody. We also demonstrate the utility of this approach for single particle electron microscopy with Fluc and two additional small membrane proteins that represent different membrane protein folds, AdiC and GlpF. These studies show that the MPER epitope provides a structurally defined, rigid docking site for antibody fragments that is transferable among diverse membrane proteins and can be engineered without prior structural information. Antibodies that bind to the MPER epitope serve as effective crystallization chaperones and electron microscopy fiducial markers, enabling structural studies of challenging small membrane proteins. | ||||||
History |
|
-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
---|
-Downloads & links
-Download
PDBx/mmCIF format | 6x58.cif.gz | 230.8 KB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb6x58.ent.gz | 185.1 KB | Display | PDB format |
PDBx/mmJSON format | 6x58.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6x58_validation.pdf.gz | 473.5 KB | Display | wwPDB validaton report |
---|---|---|---|---|
Full document | 6x58_full_validation.pdf.gz | 496.3 KB | Display | |
Data in XML | 6x58_validation.xml.gz | 40.8 KB | Display | |
Data in CIF | 6x58_validation.cif.gz | 55.3 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/x5/6x58 ftp://data.pdbj.org/pub/pdb/validation_reports/x5/6x58 | HTTPS FTP |
-Related structure data
Related structure data | 5a43S S: Starting model for refinement C: citing same article (ref.) |
---|---|
Similar structure data |
-Links
-Assembly
Deposited unit |
| ||||||||
---|---|---|---|---|---|---|---|---|---|
1 |
| ||||||||
Unit cell |
|
-Components
#1: Antibody | Mass: 25138.002 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK / Production host: Homo sapiens (human) / Strain (production host): Expi293 #2: Antibody | Mass: 22560.049 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK / Production host: Homo sapiens (human) / Strain (production host): Expi293 #3: Protein | Mass: 15383.293 Da / Num. of mol.: 2 / Mutation: R25K,A51M in transporter Source method: isolated from a genetically manipulated source Source: (gene. exp.) Human immunodeficiency virus, (gene. exp.) Escherichia coli (E. coli) Gene: env, crcB, crcB_2, flc_2, AC789_145pl00540, AKG29_01220, B6V57_25995, B7C53_24430, B9T59_30510, BANRA_05655, BANRA_05710, BEN53_26765, BIU72_24510, BK292_28205, BK334_22235, BK373_23795, BON66_ ...Gene: env, crcB, crcB_2, flc_2, AC789_145pl00540, AKG29_01220, B6V57_25995, B7C53_24430, B9T59_30510, BANRA_05655, BANRA_05710, BEN53_26765, BIU72_24510, BK292_28205, BK334_22235, BK373_23795, BON66_22385, BON69_17555, BON72_04410, BON76_23080, BON86_18360, BON94_16600, BVL39_28685, C2M16_27610, C4K41_18470, C4M78_26960, C5F73_28830, C5P01_26640, C5P44_26785, C6B13_25705, C7B08_22045, C9160_28010, C9201_28610, C9E25_24405, CDL37_21115, CSB64_25975, CWS33_26140, D1912_23150, D2184_26920, D2188_23810, D4U49_23430, D7W70_25855, D7Y10_22455, D9C99_23085, D9D33_24680, D9D94_24325, D9F87_24785, D9G11_25285, D9H70_24700, D9J44_26040, D9J61_22795, D9K02_24930, D9K48_12550, D9K54_12845, D9L99_22160, DAH18_26070, DB359_26980, DL545_01300, DLU67_24300, DLU82_24070, DN808_21420, DNI21_22025, DNQ45_14105, DNX19_24210, DP265_23725, DP277_24230, DQF72_24030, DS966_25260, DTL90_25845, DWB25_27055, E2112_25200, E2148_25435, EA239_25270, EA250_26195, EA429_25950, EA435_26495, ECONIH1_26550, ECS286_0026, EIA08_25210, EIZ93_19245, EJ366_00165, ELT22_23900, ELT58_23370, ELX56_24555, ELX56_26650, ELY05_20840, ELY05_25530, EPS76_20210, EVY14_24275, ExPECSC038_03841, EXX06_27990, EXX23_23730, EXX55_27910, EXX87_27755, EYX82_20275, F0L67_16895, F1E19_23340, F9050_24150, FORC82_p488, FQU83_00900, G3565_28150, GP698_23185, GQE58_24285, HVW93_24610, HXS78_24215, MJ49_27125, NCTC9001_00147, NCTC9077_06349, NCTC9434_05106, NCTC9969_05488, pCTXM15_EC8_00123, pO103_22, RCS79_P0115, SAMEA3472090_04449, SAMEA3752620_04806, SAMEA3753164_04868, SAMEA4370473_00090, TUM18780_48590, WP2S18E08_P10360, WP5S18E08_P10940 Production host: Escherichia coli (E. coli) / References: UniProt: Q73372, UniProt: Q6J5N4 |
---|
-Experimental details
-Experiment
Experiment | Method: X-RAY DIFFRACTION / Number of used crystals: 1 |
---|
-Sample preparation
Crystal | Density Matthews: 3.8 Å3/Da / Density % sol: 67.66 % |
---|---|
Crystal grow | Temperature: 294 K / Method: vapor diffusion, sitting drop / Details: 34-39% PEG 300, 0.1 M NaCl, 0.1 M MES pH 6.2-7 / PH range: 6.2-7 |
-Data collection
Diffraction | Mean temperature: 100 K / Serial crystal experiment: N | ||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Diffraction source | Source: SYNCHROTRON / Site: APS / Beamline: 21-ID-D / Wavelength: 0.976 Å | ||||||||||||||||||||||||||||||
Detector | Type: DECTRIS EIGER X 9M / Detector: PIXEL / Date: Jun 9, 2019 | ||||||||||||||||||||||||||||||
Radiation | Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray | ||||||||||||||||||||||||||||||
Radiation wavelength | Wavelength: 0.976 Å / Relative weight: 1 | ||||||||||||||||||||||||||||||
Reflection | Resolution: 3.26→38.18 Å / Num. obs: 28368 / % possible obs: 98.6 % / Redundancy: 10.8 % / CC1/2: 0.999 / Rmerge(I) obs: 0.135 / Rpim(I) all: 0.043 / Rrim(I) all: 0.141 / Net I/σ(I): 10.3 / Num. measured all: 306181 / Scaling rejects: 3 | ||||||||||||||||||||||||||||||
Reflection shell | Diffraction-ID: 1
|
-Processing
Software |
| ||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Refinement | Method to determine structure: MOLECULAR REPLACEMENT Starting model: 5A43 Resolution: 3.26→34.02 Å / SU ML: 0.68 / Cross valid method: THROUGHOUT / σ(F): 1.96 / Phase error: 39.74 / Stereochemistry target values: ML
| ||||||||||||||||||||
Solvent computation | Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL | ||||||||||||||||||||
Displacement parameters | Biso max: 322.54 Å2 / Biso mean: 159.7412 Å2 / Biso min: 30 Å2 | ||||||||||||||||||||
Refinement step | Cycle: final / Resolution: 3.26→34.02 Å
|