PDB-6pjx: Crystal Structure of G Protein-Coupled Receptor Kinase 5 (GRK5) i...

ID or keywords:

Entry

Database: PDB / ID: 6pjx

Title

Crystal Structure of G Protein-Coupled Receptor Kinase 5 (GRK5) in Complex with Calmodulin (CaM)

Components

Calmodulin
G protein-coupled receptor kinase 5

Keywords

TRANSFERASE / G protein-coupled receptor (GPCR) / G protein-coupled receptor kinase / phosphorylation / protein kinase / receptor regulation / signal transduction / Calmodulin / Calcium binding protein

Function / homology

Function and homology information

G-protein-coupled receptor kinase / beta-adrenergic receptor kinase activity / G protein-coupled receptor kinase activity / tachykinin receptor signaling pathway / regulation of G protein-coupled receptor signaling pathway / fat cell differentiation / regulation of signal transduction / enzyme regulator activity / protein kinase C binding / phospholipid binding ...
Similarity search - Function

Biological species

Homo sapiens (human)

Podiceps cristatus (great crested grebe)

Method

X-RAY DIFFRACTION /

MOLECULAR REPLACEMENT / Resolution: 1.96 Å

Authors

Bhardwaj, A. / Komolov, K.E. / Sulon, S. / Benovic, J.L.

Funding support

United States, 3items

Organization	Grant number	Country
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)	R01HL142310	United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)	5R35GM122541	United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)	4R01GM044944	United States

Citation

Journal: Mol.Cell / Year: 2021
Title: Structure of a GRK5-Calmodulin Complex Reveals Molecular Mechanism of GRK Activation and Substrate Targeting.
Authors: Komolov, K.E. / Sulon, S.M. / Bhardwaj, A. / van Keulen, S.C. / Duc, N.M. / Laurinavichyute, D.K. / Lou, H.J. / Turk, B.E. / Chung, K.Y. / Dror, R.O. / Benovic, J.L.

History

Deposition	Jun 28, 2019	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Dec 9, 2020	Provider: repository / Type: Initial release
Revision 1.1	Dec 30, 2020	Group: Database references / Category: citation / citation_author Item: _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2	Feb 3, 2021	Group: Database references / Category: citation Item: _citation.journal_volume / _citation.page_first / _citation.year
Revision 1.3	Oct 11, 2023	Group: Data collection / Database references / Refinement description Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession
Revision 1.4	Nov 6, 2024	Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature / Item: _pdbx_entry_details.has_protein_modification

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	6pjx.cif.gz	280.3 KB	Display	PDBx/mmCIF format
PDB format	pdb6pjx.ent.gz	222.1 KB	Display	PDB format
PDBx/mmJSON format	6pjx.json.gz		Tree view	PDBx/mmJSON format
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Validation report

Summary document	6pjx_validation.pdf.gz	815.9 KB	Display	wwPDB validaton report
Full document	6pjx_full_validation.pdf.gz	819.7 KB	Display
Data in XML	6pjx_validation.xml.gz	28.9 KB	Display
Data in CIF	6pjx_validation.cif.gz	43.2 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/pj/6pjx ftp://data.pdbj.org/pub/pdb/validation_reports/pj/6pjx	HTTPS FTP

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Related structure data

Related structure data	4tnbS 4tnd S: Starting model for refinement
Similar structure data	1v6j-d 4ik4-1 6vc4-d 6y9p-d 3evv-1 4doh-2 2ehw-d 1g6q-2 4zqb-d 4doh-1 Search similar-shape structures by Omokage search (details) Similarity search - Function & homologyF&H Search Similarity search - FunctionF&H Search Similarity search - HomologyF&H Search

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Links

PDB pages	PDBj / wwPDB / NCBI
Related items in Molecule of the Month	#44 - Aug 2003 Calmodulin similarity (1) #60 - Dec 2004 Ubiquitin similarity (5) #152 - Aug 2012 cAMP-dependent Protein Kinase similarity (3) #196 - Apr 2016 Lead Poisoning similarity (3) #183 - Mar 2015 Phototropin similarity (3)

Deposited unit

A: G protein-coupled receptor kinase 5

B: Calmodulin

hetero molecules

85.1 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

Idetical with deposited unit
defined by author&software
Evidence: isothermal titration calorimetry, gel filtration, mass spectrometry, equilibrium centrifugation, surface plasmon resonance

Unit cell

Length a, b, c (Å)	69.875, 83.061, 137.521
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	19
Space group name H-M	P2₁2₁2₁

#1: Protein	G protein-coupled receptor kinase 5 / G protein-coupled receptor kinase GRK5 Mass: 67968.711 Da / Num. of mol.: 1 / Mutation: E104K, R304H, G438E Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: GRK5, GPRK5 / Production host: Spodoptera frugiperda (fall armyworm) References: UniProt: P34947, G-protein-coupled receptor kinase
#2: Protein	Calmodulin Mass: 16721.350 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Podiceps cristatus (great crested grebe) Gene: N338_03747 / Production host: Escherichia coli (E. coli) / References: UniProt: A0A094NJQ6
#3: Chemical	ChemComp-SGV / SANGIVAMYCIN / 4-amino-7-beta-D-ribofuranosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carboxamide Mass: 309.278 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₁₂H₁₅N₅O₅
#4: Chemical	ChemComp-CA / CALCIUM ION Mass: 40.078 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Ca
#5: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 520 / Source method: isolated from a natural source / Formula: H₂O
Has ligand of interest	N
Has protein modification	Y

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Experiment

Experiment	Method: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

Crystal	Density Matthews: 2.4 Å³/Da / Density % sol: 48.9 %
Crystal grow	Temperature: 277.15 K / Method: vapor diffusion, sitting drop / pH: 7.5 Details: Sample- 25 mg/ml GRK5/CaM complex in buffer 20 mM Hepes, pH 7.2, 200 mM NaCl, 2 mM CaCl2, 1 mM DTT incubated with SGV in the presence of MgCl2. Crystallization screening reservoir condition- ...Details: Sample- 25 mg/ml GRK5/CaM complex in buffer 20 mM Hepes, pH 7.2, 200 mM NaCl, 2 mM CaCl2, 1 mM DTT incubated with SGV in the presence of MgCl2. Crystallization screening reservoir condition- 90% of (200 mM KCl, 50 mM Hepes pH 7.5, 30% 5/4 PO/OH) + 10% of (0.05% w/v L-Citrulline, 0.05% w/v Glycine, 0.05% w/v L-(-)-Threonine, 0.05% w/v L-(+)-Lysine, 0.05% w/v L-Alanine, 0.05% w/v L-Arginine, 0.05% w/v L-Asparagine monohydrate, 0.05% w/v L-Aspartic acid, 0.05% w/v L-Glutamic acid, 0.05% w/v L-Glutamine, 0.05% w/v L-Histidine, 0.05% w/v L-Isoleucine, 0.05% w/v L-Leucine, 0.05% w/v L-Methionine, 0.05% w/v L-Phenylalanine, 0.05% w/v L-Proline, 0.05% w/v L-Serine, 0.05% w/v L-Tryptophan, 0.05% w/v L-Tyrosine, 0.05% w/v L-Valine, 0.02 M HEPES sodium pH 6)

Crystal

Density Matthews: 2.4 Å³/Da / Density % sol: 48.9 %

Crystal grow

Temperature: 277.15 K / Method: vapor diffusion, sitting drop / pH: 7.5
Details: Sample- 25 mg/ml GRK5/CaM complex in buffer 20 mM Hepes, pH 7.2, 200 mM NaCl, 2 mM CaCl2, 1 mM DTT incubated with SGV in the presence of MgCl2. Crystallization screening reservoir condition- ...

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Data collection

Diffraction	Mean temperature: 93.15 K / Ambient temp details: X-stream 2000 system / Serial crystal experiment: N
Diffraction source	Source: ROTATING ANODE / Type: RIGAKU MICROMAX-007 HF / Wavelength: 1.54 Å
Detector	Type: DECTRIS PILATUS 200K / Detector: PIXEL / Date: Dec 15, 2015 / Details: Varimax-HF Confocal Optical system
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 1.54 Å / Relative weight: 1
Reflection	Resolution: 1.958→49.01 Å / Num. obs: 51717 / % possible obs: 88.5 % / Redundancy: 3.9 % / B_iso Wilson estimate: 23.97 Å² / R_merge(I) obs: 0.1077 / Net I/σ(I): 14.03
Reflection shell	Resolution: 1.96→2.03 Å / Redundancy: 2.8 % / R_merge(I) obs: 0.5964 / Mean I/σ(I) obs: 2 / % possible all: 63.7

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Processing

Software

Name	Version	Classification
PHENIX	1.15.2_3472	refinement
HKL-3000		data reduction
HKL-3000		data scaling
PHASER		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 4TNB

4tnb

Resolution: 1.96→49.01 Å / SU ML: 0.19 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 23.15

	R_factor	Num. reflection	% reflection	Selection details
R_free	0.23	3591	3.85 %	Random
R_work	0.196	-	-	-
obs	0.197	93159	83.7 %	-

Solvent computation

Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å

Displacement parameters

B_iso mean: 35.47 Å²

Refinement step

Cycle: LAST / Resolution: 1.96→49.01 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4933	0	24	520	5477

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.008	5087
X-RAY DIFFRACTION	f_angle_d	0.901	6853
X-RAY DIFFRACTION	f_dihedral_angle_d	5.47	4358
X-RAY DIFFRACTION	f_chiral_restr	0.054	728
X-RAY DIFFRACTION	f_plane_restr	0.006	898

LS refinement shell

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Refine-ID	% reflection obs (%)
1.9582-1.984	0.3605	78	0.378	2116	X-RAY DIFFRACTION	51
1.984-2.0111	0.3471	97	0.2865	2424	X-RAY DIFFRACTION	59
2.0111-2.0399	0.288	110	0.272	2656	X-RAY DIFFRACTION	65
2.0399-2.0703	0.2545	117	0.259	2898	X-RAY DIFFRACTION	71
2.0703-2.1027	0.2625	126	0.2467	3173	X-RAY DIFFRACTION	77
2.1027-2.1372	0.2408	144	0.2375	3572	X-RAY DIFFRACTION	86
2.1372-2.174	0.2135	150	0.2266	3650	X-RAY DIFFRACTION	89
2.174-2.2135	0.2479	143	0.2248	3636	X-RAY DIFFRACTION	89
2.2135-2.2561	0.2596	149	0.2198	3704	X-RAY DIFFRACTION	89
2.2561-2.3022	0.2806	148	0.2205	3624	X-RAY DIFFRACTION	89
2.3022-2.3522	0.2331	151	0.2168	3657	X-RAY DIFFRACTION	89
2.3522-2.4069	0.2735	145	0.2126	3677	X-RAY DIFFRACTION	89
2.4069-2.4671	0.2653	149	0.2138	3704	X-RAY DIFFRACTION	90
2.4671-2.5338	0.2612	149	0.212	3718	X-RAY DIFFRACTION	90
2.5338-2.6084	0.2659	152	0.2208	3765	X-RAY DIFFRACTION	92
2.6084-2.6926	0.2713	155	0.216	3883	X-RAY DIFFRACTION	93
2.6926-2.7888	0.268	149	0.2089	3828	X-RAY DIFFRACTION	93
2.7888-2.9004	0.3026	152	0.2077	3745	X-RAY DIFFRACTION	92
2.9004-3.0324	0.2088	155	0.2098	3723	X-RAY DIFFRACTION	90
3.0324-3.1923	0.2282	147	0.2002	3629	X-RAY DIFFRACTION	88
3.1923-3.3923	0.1912	135	0.1887	3531	X-RAY DIFFRACTION	86
3.3923-3.6541	0.1854	138	0.1691	3409	X-RAY DIFFRACTION	83
3.6541-4.0217	0.2238	136	0.1615	3385	X-RAY DIFFRACTION	82
4.0217-4.6033	0.1599	135	0.1503	3404	X-RAY DIFFRACTION	83
4.6033-5.7982	0.2246	143	0.1571	3497	X-RAY DIFFRACTION	85
5.7982-49.8523	0.2042	138	0.1662	3560	X-RAY DIFFRACTION	86

Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

ID	L11 (°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)
1	3.1921	3.0293	1.1365	2.8519	0.9068	0.6896	-0.124	0.2948	-0.3492	-0.017	0.2657	-0.2686	0.082	0.0304	-0.1727	0.1918	0.0373	0.039	0.1848	-0.0356	0.1974	-0.5801	-5.3366	-24.8311
2	3.2543	-1.2062	0.7435	3.2886	-0.7987	3.642	0.1213	0.4874	0.7348	-0.3983	-0.1231	0.1564	-0.6451	-0.373	-0.0169	0.3346	0.0923	0.0107	0.311	0.0568	0.3905	-28.9792	8.0173	-41.2337
3	0.9091	0.3572	0.2679	1.2228	0.2514	0.601	0.0642	-0.1563	-0.0121	0.1951	-0.0932	-0.0493	0.0024	-0.0639	0.0553	0.1496	-0.0128	0.0326	0.1536	-0.0228	0.0988	-8.5399	1.7592	-15.7684
4	3.1408	0.0046	-1.3221	1.4798	0.1149	2.5012	0.1496	0.1023	0.2733	-0.0362	-0.0578	0.0581	-0.3137	-0.2494	-0.1037	0.1794	0.0473	0.0273	0.1457	0.0021	0.1084	-3.3109	25.6066	-21.2775
5	3.0231	-0.3413	-2.278	1.7575	0.4582	3.8758	0.44	-0.0128	0.6334	0.0142	-0.0182	0.0363	-0.7687	-0.1431	-0.1691	0.3036	0.0191	0.1051	0.1517	-0.0193	0.25	-7.8145	31.563	-12.9445
6	1.5757	0.318	0.3146	0.9411	0.8805	1.6416	0.1988	-0.392	-0.1852	0.3105	-0.2067	-0.079	0.4031	-0.1445	0.0974	0.2884	-0.0801	-0.012	0.2536	0.0347	0.1219	2.3708	7.5392	-5.971
7	1.6373	1.0868	0.7061	3.9446	3.4818	4.3946	-0.0094	0.0009	-0.133	0.095	-0.0112	0.0155	0.2464	0.0261	0.034	0.1255	0.0088	0.0164	0.1153	-0.007	0.1298	-12.4513	-9.8132	-30.2816
8	8.5858	3.179	-4.362	3.903	-4.0043	4.4876	-0.6517	-0.5079	-0.9267	0.1857	0.3767	0.5648	0.6026	-0.4537	0.1596	0.3454	0.086	0.1486	0.2947	0.0263	0.4497	17.1333	-4.1511	-19.1038
9	7.319	3.9942	0.5366	4.4549	4.7064	8.7097	0.0425	0.472	0.4193	-0.0853	-0.0265	0.1437	-0.0324	0.09	-0.0752	0.1261	0.0492	0.0071	0.1905	-0.0293	0.2965	28.0862	-0.3371	-28.0058
10	0.9898	0.7991	0.2086	1.5903	0.7435	0.4676	0.3231	0.5674	0.7838	-0.8879	-0.0178	-1.2268	-0.3177	0.1286	-0.2164	0.2544	0.0495	0.1283	0.2122	0.1159	0.4639	24.6361	12.7428	-28.4576
11	4.7017	0.5366	-0.2682	4.7654	1.5094	2.8498	-0.1094	-0.7466	0.6898	0.6134	0.4284	-0.5896	0.1942	0.6238	-0.1775	0.2037	0.109	-0.049	0.3907	-0.1167	0.4212	29.6042	5.3641	-17.8614

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	CHAIN 'A' AND (RESID 2 THROUGH 48 )
2	X-RAY DIFFRACTION	2	CHAIN 'A' AND (RESID 49 THROUGH 153 )
3	X-RAY DIFFRACTION	3	CHAIN 'A' AND (RESID 154 THROUGH 263 )
4	X-RAY DIFFRACTION	4	CHAIN 'A' AND (RESID 264 THROUGH 305 )
5	X-RAY DIFFRACTION	5	CHAIN 'A' AND (RESID 306 THROUGH 467 )
6	X-RAY DIFFRACTION	6	CHAIN 'A' AND (RESID 468 THROUGH 502 )
7	X-RAY DIFFRACTION	7	CHAIN 'A' AND (RESID 503 THROUGH 542 )
8	X-RAY DIFFRACTION	8	CHAIN 'B' AND (RESID 4 THROUGH 19 )
9	X-RAY DIFFRACTION	9	CHAIN 'B' AND (RESID 20 THROUGH 28 )
10	X-RAY DIFFRACTION	10	CHAIN 'B' AND (RESID 29 THROUGH 44 )
11	X-RAY DIFFRACTION	11	CHAIN 'B' AND (RESID 45 THROUGH 73 )

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