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- EMDB-23977: Rhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin -

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Basic information

Entry
Database: EMDB / ID: EMD-23977
TitleRhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin
Map dataRhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin
Sample
  • Complex: Rhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin
    • Protein or peptide: Rhodopsin kinase GRK1
    • Protein or peptide: Rhodopsin
  • Ligand: SANGIVAMYCIN
  • Ligand: RETINAL
Function / homology
Function and homology information


rhodopsin kinase / rhodopsin kinase activity / The canonical retinoid cycle in rods (twilight vision) / Opsins / VxPx cargo-targeting to cilium / regulation of rhodopsin mediated signaling pathway / rod photoreceptor outer segment / G protein-coupled receptor complex / absorption of visible light / opsin binding ...rhodopsin kinase / rhodopsin kinase activity / The canonical retinoid cycle in rods (twilight vision) / Opsins / VxPx cargo-targeting to cilium / regulation of rhodopsin mediated signaling pathway / rod photoreceptor outer segment / G protein-coupled receptor complex / absorption of visible light / opsin binding / G protein-coupled photoreceptor activity / photoreceptor inner segment membrane / 11-cis retinal binding / rhodopsin mediated signaling pathway / cellular response to light stimulus / sperm head plasma membrane / Inactivation, recovery and regulation of the phototransduction cascade / arrestin family protein binding / phototransduction, visible light / Activation of the phototransduction cascade / thermotaxis / detection of temperature stimulus involved in thermoception / outer membrane / photoreceptor cell maintenance / photoreceptor outer segment membrane / G alpha (i) signalling events / G-protein alpha-subunit binding / response to light stimulus / photoreceptor outer segment / phototransduction / regulation of signal transduction / sperm midpiece / visual perception / photoreceptor disc membrane / guanyl-nucleotide exchange factor activity / retina development in camera-type eye / cell-cell junction / G protein-coupled receptor signaling pathway / protein kinase activity / protein autophosphorylation / Golgi membrane / protein phosphorylation / signal transduction / zinc ion binding / ATP binding / membrane / identical protein binding / plasma membrane / cytoplasm
Similarity search - Function
Rhodopsin kinase GRK1 / Rhodopsin kinase, catalytic domain / GPCR kinase / Rhodopsin, N-terminal / Amino terminal of the G-protein receptor rhodopsin / Rhodopsin / Opsin / Visual pigments (opsins) retinal binding site / Visual pigments (opsins) retinal binding site. / Regulator of G protein signaling domain ...Rhodopsin kinase GRK1 / Rhodopsin kinase, catalytic domain / GPCR kinase / Rhodopsin, N-terminal / Amino terminal of the G-protein receptor rhodopsin / Rhodopsin / Opsin / Visual pigments (opsins) retinal binding site / Visual pigments (opsins) retinal binding site. / Regulator of G protein signaling domain / RGS, subdomain 2 / RGS domain / RGS domain profile. / Regulator of G protein signalling domain / RGS domain superfamily / Serpentine type 7TM GPCR chemoreceptor Srsx / Extension to Ser/Thr-type protein kinases / AGC-kinase, C-terminal / AGC-kinase C-terminal domain profile. / G-protein coupled receptors family 1 signature. / 7 transmembrane receptor (rhodopsin family) / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. / Protein kinase domain / Serine/Threonine protein kinases, catalytic domain / Protein kinase, ATP binding site / Protein kinases ATP-binding region signature. / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Rhodopsin / Rhodopsin kinase GRK1
Similarity search - Component
Biological speciesBos taurus (cattle) / Bovine (cattle)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.8 Å
AuthorsChen Q / Li Z / Chang L / Tesmer JJG
Funding support United States, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)HL071818 United States
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)HL122416 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA221289 United States
American Heart Association19POST34450193 United States
CitationJournal: Nature / Year: 2021
Title: Structures of rhodopsin in complex with G-protein-coupled receptor kinase 1.
Authors: Qiuyan Chen / Manolo Plasencia / Zhuang Li / Somnath Mukherjee / Dhabaleswar Patra / Chun-Liang Chen / Thomas Klose / Xin-Qiu Yao / Anthony A Kossiakoff / Leifu Chang / Philip C Andrews / John J G Tesmer /
Abstract: G-protein-coupled receptor (GPCR) kinases (GRKs) selectively phosphorylate activated GPCRs, thereby priming them for desensitization. Although it is unclear how GRKs recognize these receptors, a ...G-protein-coupled receptor (GPCR) kinases (GRKs) selectively phosphorylate activated GPCRs, thereby priming them for desensitization. Although it is unclear how GRKs recognize these receptors, a conserved region at the GRK N terminus is essential for this process. Here we report a series of cryo-electron microscopy single-particle reconstructions of light-activated rhodopsin (Rho*) bound to rhodopsin kinase (GRK1), wherein the N terminus of GRK1 forms a helix that docks into the open cytoplasmic cleft of Rho*. The helix also packs against the GRK1 kinase domain and stabilizes it in an active configuration. The complex is further stabilized by electrostatic interactions between basic residues that are conserved in most GPCRs and acidic residues that are conserved in GRKs. We did not observe any density for the regulator of G-protein signalling homology domain of GRK1 or the C terminus of rhodopsin. Crosslinking with mass spectrometry analysis confirmed these results and revealed dynamic behaviour in receptor-bound GRK1 that would allow the phosphorylation of multiple sites in the receptor tail. We have identified GRK1 residues whose mutation augments kinase activity and crosslinking with Rho*, as well as residues that are involved in activation by acidic phospholipids. From these data, we present a general model for how a small family of protein kinases can recognize and be activated by hundreds of different GPCRs.
History
DepositionMay 13, 2021-
Header (metadata) releaseJul 7, 2021-
Map releaseJul 7, 2021-
UpdateAug 25, 2021-
Current statusAug 25, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.062
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.062
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7mt8
  • Surface level: 0.062
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7mt8
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_23977.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationRhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 256 pix.
= 276.48 Å
1.08 Å/pix.
x 256 pix.
= 276.48 Å
1.08 Å/pix.
x 256 pix.
= 276.48 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.08 Å
Density
Contour LevelBy AUTHOR: 0.059 / Movie #1: 0.062
Minimum - Maximum-0.17262281 - 0.29358274
Average (Standard dev.)0.00047108135 (±0.0061750943)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 276.48 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.081.081.08
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z276.480276.480276.480
α/β/γ90.00090.00090.000
start NX/NY/NZ-210-210-210
NX/NY/NZ420420420
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.1730.2940.000

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Supplemental data

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Sample components

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Entire : Rhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin

EntireName: Rhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin
Components
  • Complex: Rhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin
    • Protein or peptide: Rhodopsin kinase GRK1
    • Protein or peptide: Rhodopsin
  • Ligand: SANGIVAMYCIN
  • Ligand: RETINAL

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Supramolecule #1: Rhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin

SupramoleculeName: Rhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Bos taurus (cattle)

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Macromolecule #1: Rhodopsin kinase GRK1

MacromoleculeName: Rhodopsin kinase GRK1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: rhodopsin kinase
Source (natural)Organism: Bos taurus (cattle)
Molecular weightTheoretical: 61.542012 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MDFGELETVV ANSAFIAARG SFDASSGPAS RDRKYLARLK LPPLSKCEAL RESLDLGFEG MCLEQPIGKR LFQQFLRTHE QHGPALQLW KDIEDYDTAD DALRPQKAQA LRAAYLEPQA QLFCSFLDAE TVARARAGAG DGLFQPLLRA VLAHLGQAPF Q EFLDSLYF ...String:
MDFGELETVV ANSAFIAARG SFDASSGPAS RDRKYLARLK LPPLSKCEAL RESLDLGFEG MCLEQPIGKR LFQQFLRTHE QHGPALQLW KDIEDYDTAD DALRPQKAQA LRAAYLEPQA QLFCSFLDAE TVARARAGAG DGLFQPLLRA VLAHLGQAPF Q EFLDSLYF LRFLQWKWLE AQPMGEDWFL DFRVLGRGGF GEVFACQMKA TGKLYACKKL NKKRLKKRKG YQGAMVEKKI LA KVHSRFI VSLAYAFETK TDLCLVMTIM NGGDIRYHIY NVDEDNPGFQ EPRAIFYTAQ IVSGLEHLHQ RNIIYRDLKP ENV LLDDDG NVRISDLGLA VELKAGQTKT KGYAGTPGFM APELLLGEEY DFSVDYFALG VTLYEMIAAR GPFRARGEKV ENKE LKQRV LEQAVTYPDK FSPASKDFCE ALLQKDPEKR LGFRDGSCDG LRTHPLFRDI SWRQLEAGML TPPFVPDSRT VYAKN IQDV GAFEEVKGVA FEKADTEFFQ EFASGTCPIP WQEEMIETGV FGDLNVWRPD GVDHHHHHH

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Macromolecule #2: Rhodopsin

MacromoleculeName: Rhodopsin / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bovine (cattle)
Molecular weightTheoretical: 39.031457 KDa
SequenceString: MNGTEGPNFY VPFSNKTGVV RSPFEAPQYY LAEPWQFSML AAYMFLLIML GFPINFLTLY VTVQHKKLRT PLNYILLNLA VADLFMVFG GFTTTLYTSL HGYFVFGPTG CNLEGFFATL GGEIALWSLV VLAIERYVVV CKPMSNFRFG ENHAIMGVAF T WVMALACA ...String:
MNGTEGPNFY VPFSNKTGVV RSPFEAPQYY LAEPWQFSML AAYMFLLIML GFPINFLTLY VTVQHKKLRT PLNYILLNLA VADLFMVFG GFTTTLYTSL HGYFVFGPTG CNLEGFFATL GGEIALWSLV VLAIERYVVV CKPMSNFRFG ENHAIMGVAF T WVMALACA APPLVGWSRY IPEGMQCSCG IDYYTPHEET NNESFVIYMF VVHFIIPLIV IFFCYGQLVF TVKEAAAQQQ ES ATTQKAE KEVTRMVIIM VIAFLICWLP YAGVAFYIFT HQGSDFGPIF MTIPAFFAKT SAVYNPVIYI MMNKQFRNCM VTT LCCGKN PLGDDEASTT VSKTETSQVA PA

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Macromolecule #3: SANGIVAMYCIN

MacromoleculeName: SANGIVAMYCIN / type: ligand / ID: 3 / Number of copies: 1 / Formula: SGV
Molecular weightTheoretical: 309.278 Da
Chemical component information

ChemComp-SGV:
SANGIVAMYCIN / inhibitor*YM / Sangivamycin

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Macromolecule #4: RETINAL

MacromoleculeName: RETINAL / type: ligand / ID: 4 / Number of copies: 1 / Formula: RET
Molecular weightTheoretical: 284.436 Da
Chemical component information

ChemComp-RET:
RETINAL / Retinal

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 54.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 5.8 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 132721
FSC plot (resolution estimation)

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