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- PDB-7mta: Rhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin... -

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Basic information

Entry
Database: PDB / ID: 7mta
TitleRhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin and Fab1
Components
  • Fab1 Heavy chainPIKFYVE
  • Fab1 Light chainPIKFYVE
  • Rhodopsin kinase GRK1
  • Rhodopsin
KeywordsMEMBRANE / Signaling protein/Immune System / GPCR / signal desensitization / Kinase / MEMBRANE PROTEIN / Signaling protein-Immune System complex
Function / homology
Function and homology information


rhodopsin kinase / rhodopsin kinase activity / regulation of rhodopsin mediated signaling pathway / Opsins / VxPx cargo-targeting to cilium / rod photoreceptor outer segment / rod bipolar cell differentiation / sperm head plasma membrane / podosome assembly / absorption of visible light ...rhodopsin kinase / rhodopsin kinase activity / regulation of rhodopsin mediated signaling pathway / Opsins / VxPx cargo-targeting to cilium / rod photoreceptor outer segment / rod bipolar cell differentiation / sperm head plasma membrane / podosome assembly / absorption of visible light / opsin binding / The canonical retinoid cycle in rods (twilight vision) / adaptation of rhodopsin mediated signaling / G protein-coupled photoreceptor activity / photoreceptor inner segment membrane / rhodopsin mediated signaling pathway / 11-cis retinal binding / cellular response to light stimulus / G protein-coupled receptor complex / Inactivation, recovery and regulation of the phototransduction cascade / phototransduction, visible light / Activation of the phototransduction cascade / thermotaxis / detection of temperature stimulus involved in thermoception / outer membrane / arrestin family protein binding / photoreceptor cell maintenance / photoreceptor outer segment membrane / G alpha (i) signalling events / response to light stimulus / phototransduction / photoreceptor outer segment / G-protein alpha-subunit binding / regulation of signal transduction / sperm midpiece / visual perception / guanyl-nucleotide exchange factor activity / microtubule cytoskeleton organization / photoreceptor disc membrane / cell-cell junction / gene expression / protein autophosphorylation / G protein-coupled receptor signaling pathway / Golgi membrane / signal transduction / zinc ion binding / ATP binding / membrane / identical protein binding / plasma membrane / cytoplasm
Similarity search - Function
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 ...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 / Extension to Ser/Thr-type protein kinases / AGC-kinase, C-terminal / AGC-kinase C-terminal domain profile. / Serpentine type 7TM GPCR chemoreceptor Srsx / G-protein coupled receptors family 1 signature. / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / 7 transmembrane receptor (rhodopsin family) / 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
RETINAL / SANGIVAMYCIN / Rhodopsin / Rhodopsin kinase GRK1
Similarity search - Component
Biological speciesBos taurus (cattle)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsChen, Q. / Chen, C.-L. / Tesmer, J.J.G.
Funding support United States, 4items
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, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 7, 2021Provider: repository / Type: Initial release
Revision 1.1Jul 28, 2021Group: Database references / Category: 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_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Aug 25, 2021Group: Database references / Category: citation / citation_author / database_2
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID / _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Assembly

Deposited unit
G: Rhodopsin kinase GRK1
H: Fab1 Heavy chain
L: Fab1 Light chain
R: Rhodopsin
hetero molecules


Theoretical massNumber of molelcules
Total (without water)149,9536
Polymers149,3604
Non-polymers5942
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: cross-linking
TypeNameSymmetry operationNumber
identity operation1_5551
Number of models6

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Components

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Protein , 2 types, 2 molecules GR

#1: Protein Rhodopsin kinase GRK1 / RK / G protein-coupled receptor kinase 1


Mass: 61542.012 Da / Num. of mol.: 1 / Mutation: S5E, S488E, T489E
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bos taurus (cattle) / Gene: GRK1, RHOK / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P28327, rhodopsin kinase
#4: Protein Rhodopsin /


Mass: 39031.457 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Bos taurus (cattle) / References: UniProt: P02699

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Antibody , 2 types, 2 molecules HL

#2: Antibody Fab1 Heavy chain / PIKFYVE


Mass: 24999.779 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)
#3: Antibody Fab1 Light chain / PIKFYVE


Mass: 23786.396 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)

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Non-polymers , 2 types, 2 molecules

#5: Chemical ChemComp-SGV / SANGIVAMYCIN / 4-amino-7-beta-D-ribofuranosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carboxamide / Sangivamycin


Mass: 309.278 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C12H15N5O5 / Comment: inhibitor*YM
#6: Chemical ChemComp-RET / RETINAL / Retinal


Mass: 284.436 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C20H28O

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Details

Has ligand of interestN

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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

Component
IDNameTypeEntity IDParent-IDSource
1Rhodopsin kinase (GRK1)-S5E/S488E/T489E in complex with rhodopsin stabilized by Fab1COMPLEX#1-#40MULTIPLE SOURCES
2Fab Heavy and Light ChainsCOMPLEX#2-#31RECOMBINANT
3Rhodopsin kinase, rhodopsinCOMPLEX1MULTIPLE SOURCES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
13Bos taurus (cattle)9913
22Homo sapiens (human)9606
Buffer solutionpH: 8
SpecimenEmbedding 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: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 54 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 310363 / Symmetry type: POINT

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