PDB-6t8m: Prolyl Hydroxylase (PHD) involved in hypoxia sensing by Dictyoste...

ID or keywords:

Entry

Database: PDB / ID: 6t8m

Title

Prolyl Hydroxylase (PHD) involved in hypoxia sensing by Dictyostelium discoideum

Components

Prolyl 4-hydroxylase subunit alpha

Keywords

OXIDOREDUCTASE / Prolyl Hydroxylase / Oxygen Sensing / 2-oxoglutarate and iron dependent oxygenase

Function / homology

Function and homology information

oxygen metabolic process / SCF complex assembly / peptidyl-proline 4-dioxygenase activity / culmination involved in sorocarp development / peptidyl-proline dioxygenase activity / peptidyl-proline hydroxylation to 4-hydroxy-L-proline / Oxidoreductases; Acting on paired donors, with incorporation or reduction of molecular oxygen; With 2-oxoglutarate as one donor, and incorporation of one atom of oxygen into each donor / L-ascorbic acid binding / ferrous iron binding / cellular response to hypoxia / cytoplasm
Similarity search - Function

Biological species

Dictyostelium discoideum (eukaryote)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.02 Å

Authors

Chowdhury, R. / McDonough, M.A. / Liu, T. / Clifton, I.J. / Schofield, C.J.

Funding support

United Kingdom, 1items

Citation

Journal: J.Biol.Chem. / Year: 2020
Title: Biochemical and biophysical analyses of hypoxia sensing prolyl hydroxylases from Dictyostelium discoideum and Toxoplasma gondii .
Authors: Liu, T. / Abboud, M.I. / Chowdhury, R. / Tumber, A. / Hardy, A.P. / Lippl, K. / Lohans, C.T. / Pires, E. / Wickens, J. / McDonough, M.A. / West, C.M. / Schofield, C.J.

History

Deposition	Oct 24, 2019	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Sep 23, 2020	Provider: repository / Type: Initial release
Revision 1.1	Sep 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	Dec 16, 2020	Group: Database references / Category: citation / citation_author Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.title / _citation_author.identifier_ORCID
Revision 1.3	Jan 24, 2024	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

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	6t8m.cif.gz	290.5 KB	Display	PDBx/mmCIF format
PDB format	pdb6t8m.ent.gz	223.8 KB	Display	PDB format
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Validation report

Summary document	6t8m_validation.pdf.gz	1.4 MB	Display	wwPDB validaton report
Full document	6t8m_full_validation.pdf.gz	1.4 MB	Display
Data in XML	6t8m_validation.xml.gz	25.3 KB	Display
Data in CIF	6t8m_validation.cif.gz	34.4 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/t8/6t8m ftp://data.pdbj.org/pub/pdb/validation_reports/t8/6t8m	HTTPS FTP

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Related structure data

Related structure data	4j25S 5l9rS S: Starting model for refinement
Similar structure data	7bft-d 3oru-d 2wfm-2 3gzj-3 2wfm-3 5zdb-1 3gzj-5 6f0w-d 3c5n-2 2cbg-d Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Similarity search - Function & homologyF&H Search Similarity search - FunctionF&H Search Similarity search - HomologyF&H Search

Related structure data

4j25S

5l9rS

S: Starting model for refinement

Similar structure data

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Links

PDB pages	PDBj / wwPDB / NCBI
Related items in Molecule of the Month	#240 - Dec 2019 Hypoxia-Inducible Factors similarity (6)

Deposited unit

A: Prolyl 4-hydroxylase subunit alpha

B: Prolyl 4-hydroxylase subunit alpha

C: Prolyl 4-hydroxylase subunit alpha

hetero molecules

82 kDa, 3 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: Prolyl 4-hydroxylase subunit alpha

hetero molecules

defined by author
Evidence: gel filtration
27.3 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

B: Prolyl 4-hydroxylase subunit alpha

hetero molecules

defined by author
27.3 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

3

C: Prolyl 4-hydroxylase subunit alpha

hetero molecules

defined by author
27.4 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

Unit cell

Length a, b, c (Å)	45.216, 81.781, 379.210
Angle α, β, γ (deg.)	90.000, 90.000, 90.000
Int Tables number	20
Space group name H-M	C222₁
Space group name Hall	C2c2

Components on special symmetry positions

ID	Model	Components
1	1	C-305- NI
2	1	A-647- HOH

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Protein , 1 types, 3 molecules ABC

#1: Protein	Prolyl 4-hydroxylase subunit alpha / Prolyl 4-hydrolase Mass: 26973.525 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Dictyostelium discoideum (eukaryote) / Gene: phyA, p4h1, DDB_G0277759 / Plasmid: pNIC-ZB / Cell line (production host): BL21 GOLD / Production host: Escherichia coli (E. coli) / Variant (production host): DE3 References: UniProt: Q86KR9, Oxidoreductases; Acting on paired donors, with incorporation or reduction of molecular oxygen; With 2-oxoglutarate as one donor, and incorporation of one atom of oxygen into each donor

#1: Protein

Prolyl 4-hydroxylase subunit alpha / Prolyl 4-hydrolase

Mass: 26973.525 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.)

Dictyostelium discoideum (eukaryote) / Gene: phyA, p4h1, DDB_G0277759 / Plasmid: pNIC-ZB / Cell line (production host): BL21 GOLD / Production host:

Escherichia coli (E. coli) / Variant (production host): DE3
References: UniProt: Q86KR9, Oxidoreductases; Acting on paired donors, with incorporation or reduction of molecular oxygen; With 2-oxoglutarate as one donor, and incorporation of one atom of oxygen into each donor

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Non-polymers , 5 types, 161 molecules

#2: Chemical	ChemComp-NI / NICKEL (II) ION Mass: 58.693 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: Ni
#3: Chemical	ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL Mass: 92.094 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C₃H₈O₃
#4: Chemical	ChemComp-OGA / N-OXALYLGLYCINE Mass: 147.086 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C₄H₅NO₅ / Feature type: SUBJECT OF INVESTIGATION / Comment: inhibitor*YM
#5: Chemical	ChemComp-CL / CHLORIDE ION Mass: 35.453 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Cl
#6: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 149 / Source method: isolated from a natural source / Formula: H₂O

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Details

Has ligand of interest	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: 2.29 Å³/Da / Density % sol: 46.3 % / Description: rhombohedron
Crystal grow	Temperature: 291 K / Method: vapor diffusion, sitting drop / pH: 7.5 Details: 0.1 M HEPES pH 7.5, 17.5-23% PEG 6000, 0.2M NaCl, 2mM NiCl2, 0-6% glycerol PH range: 7.0-8.0

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

Diamond

/ Beamline: I03 / Wavelength: 0.97623 Å

Detector

Type: DECTRIS PILATUS 6M-F / Detector: PIXEL / Date: Jun 29, 2017

Radiation

Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray

Radiation wavelength

Wavelength: 0.97623 Å / Relative weight: 1

Reflection

Resolution: 2.02→189.535 Å / Num. all: 47025 / Num. obs: 47025 / % possible obs: 100 % / Redundancy: 9.1 % / B_iso Wilson estimate: 39.8 Å² / R_pim(I) all: 0.036 / R_rim(I) all: 0.11 / R_sym value: 0.103 / Net I/av σ(I): 5.9 / Net I/σ(I): 12.2 / Num. measured all: 429431

Reflection shell

Diffraction-ID: 1

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Mean I/σ(I) obs	Num. measured all	Num. unique obs	R_pim(I) all	R_rim(I) all	R_sym value	Net I/σ(I) obs	% possible all
2.02-2.13	9.1	1.426	0.5	61814	6792	0.495	1.512	1.426	1.7	100
2.13-2.26	9.2	0.93	0.8	58775	6397	0.319	0.985	0.93	2.6	100
2.26-2.41	9.5	0.652	1.2	56898	5996	0.218	0.688	0.652	3.6	99.9
2.41-2.61	9.1	0.389	1.9	51314	5644	0.133	0.412	0.389	5.3	100
2.61-2.86	9.3	0.246	3.1	48390	5178	0.083	0.26	0.246	7.9	100
2.86-3.19	9.6	0.137	5.4	45311	4742	0.046	0.145	0.137	13.4	99.9
3.19-3.69	9	0.082	8.7	37760	4190	0.028	0.087	0.082	22.1	100
3.69-4.52	8.8	0.054	12.1	31718	3603	0.019	0.057	0.054	33.5	100
4.52-6.39	8.6	0.044	14.3	24196	2817	0.016	0.047	0.044	38.2	99.9
6.39-42.119	8	0.037	13.9	13255	1666	0.014	0.04	0.037	40.7	99.8

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Processing

Software

Name	Version	Classification
PHENIX	1.14_3260	refinement
SCALA	3.3.22	data scaling
PDB_EXTRACT	3.25	data extraction
XDS		data reduction
PHASER		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 4J25, 5L9R

4j25

5l9r

Resolution: 2.02→37.92 Å / SU ML: 0.3177 / Cross valid method: THROUGHOUT / σ(F): 1.36 / Phase error: 28.9968 / Stereochemistry target values: CDL v1.2

	R_factor	Num. reflection	% reflection
R_free	0.2565	2327	4.96 %
R_work	0.2277	44635	-
obs	0.2291	46962	99.96 %

Solvent computation

Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL

Displacement parameters

B_iso mean: 60.45 Å²

Refinement step

Cycle: LAST / Resolution: 2.02→37.92 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	5058	0	54	149	5261

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.0023	5200
X-RAY DIFFRACTION	f_angle_d	0.4549	7003
X-RAY DIFFRACTION	f_chiral_restr	0.0429	758
X-RAY DIFFRACTION	f_plane_restr	0.0023	888
X-RAY DIFFRACTION	f_dihedral_angle_d	11.3887	3107

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 (%)
2.02-2.06	0.4054	124	0.4084	2602	X-RAY DIFFRACTION	100
2.06-2.11	0.3874	144	0.3977	2553	X-RAY DIFFRACTION	99.93
2.11-2.16	0.3434	148	0.3603	2592	X-RAY DIFFRACTION	99.96
2.16-2.21	0.2997	142	0.3507	2573	X-RAY DIFFRACTION	99.96
2.21-2.27	0.3199	138	0.3244	2612	X-RAY DIFFRACTION	99.93
2.27-2.34	0.3328	128	0.3131	2619	X-RAY DIFFRACTION	99.96
2.34-2.41	0.2783	104	0.2998	2570	X-RAY DIFFRACTION	99.96
2.41-2.5	0.3302	130	0.2789	2623	X-RAY DIFFRACTION	99.96
2.5-2.6	0.2898	143	0.2713	2621	X-RAY DIFFRACTION	100
2.6-2.72	0.2686	139	0.2644	2576	X-RAY DIFFRACTION	100
2.72-2.86	0.2786	148	0.2682	2595	X-RAY DIFFRACTION	99.93
2.86-3.04	0.2713	147	0.2443	2636	X-RAY DIFFRACTION	99.96
3.04-3.27	0.2636	125	0.2304	2650	X-RAY DIFFRACTION	100
3.27-3.6	0.2387	166	0.2092	2622	X-RAY DIFFRACTION	100
3.6-4.12	0.2295	124	0.185	2657	X-RAY DIFFRACTION	100
4.12-5.19	0.167	117	0.1549	2735	X-RAY DIFFRACTION	99.96
5.19-37.92	0.258	160	0.2044	2799	X-RAY DIFFRACTION	99.87

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	0.494941678906	0.23862141811	0.475279403785	0.233911535953	-0.0313252858909	0.992419756649	-0.3175248138	-0.367359466152	-1.06120765911	-0.223428137091	-0.183004990087	0.259904027959	0.728458342242	1.06631144534	0.527894257946	1.01650375737	0.298934221061	0.108926440244	0.629181703635	0.235131518081	0.878968747515	-21.1448483512	-18.5562145437	29.9925067652
2	6.15074209267	-0.459554463813	2.88455881096	5.6824582782	-1.0510976274	5.8972370501	0.167241982341	0.485607575734	-0.069338785417	0.06687645559	0.0138158164171	-0.0502608740464	0.128829280443	0.720696910829	-0.0304031264162	0.981557246464	0.276468405323	0.131667339016	0.277635231199	0.0894093379518	0.580240072949	-21.1100127352	-10.4784047549	20.9590496535
3	3.03369083858	0.196148267424	0.592128453152	0.272904402277	0.998870933534	3.58339863851	-0.219745178087	0.0563770822228	0.195987355401	-0.133859579438	0.0905317871727	0.0152288424121	-0.0107250619867	-0.0612441243476	0.135546737861	1.04295294187	0.0394180948257	0.060657720638	0.33482820677	0.0433655903923	0.398091320122	-29.1579409678	-2.14052702241	16.1411121229
4	3.05880396205	0.429579455457	0.862655158683	0.987320930759	0.189908066972	5.30184203953	-0.132945194123	0.0739365782329	-0.140911831559	-0.716437578923	0.0889768965037	0.0704405668713	0.267367055583	-0.827099536404	0.0176676770746	0.906518703102	-0.041293169118	-0.00648005447754	0.32419545659	0.103136792648	0.399056868714	-35.8455114666	-8.59235694338	17.8985113525
5	1.39044583652	-0.592779689364	-0.264107438895	1.04463715426	-0.383452259256	1.1947912076	-0.192484607619	-0.0975066184897	-0.210582810469	-0.488806877524	0.0298738726495	-0.18510236117	0.709895402777	-0.326481822139	-0.497286899314	1.39570670521	-0.0121637369376	0.000848394343051	0.181944176675	0.197208447257	0.425302804151	-33.0481717229	-11.0768534681	19.6019949075
6	3.27068181894	-1.07889488568	1.59815082637	6.89613878509	-3.30400202919	2.46134616963	0.219505068787	0.489681053125	-0.210874596604	-0.631629151441	-0.102562984695	0.427296542875	1.00280947368	-1.08070689307	-0.223353133765	0.545969853636	-0.0359361445706	-0.0532721274202	0.7607409402	0.0850814177753	0.450279648692	-34.5350242001	-13.6903111757	73.3207026805
7	2.77916370956	-0.157096230359	1.79221438294	1.84403054396	0.314987482195	1.75313673224	0.211433730539	0.658619721457	0.366978716112	-0.202516724742	-0.12378067248	-0.169881989809	0.384938885695	1.36506311573	-0.192200240878	0.318418652504	0.0814893705432	0.082782397868	0.819236697222	0.0525396636714	0.454584443871	-14.0191785709	-4.55465000512	70.0135603019
8	0.377408335433	0.0758126462664	0.820603925409	0.192368229304	0.332195747306	1.94291506558	0.341818107415	-0.0362436635629	0.0605892911282	0.0288637851843	-0.857516194148	-0.463850160425	-0.581250183695	2.2820171227	0.537716054755	0.431960821673	-0.101764455063	-0.033518393884	1.12982118722	0.128714982907	0.75819054978	-6.55764329496	-2.4240527095	92.7726099971
9	6.81394676637	0.190268513499	1.17649432549	3.99100702928	0.715818347181	6.51803723599	-0.457356756555	0.304453627007	0.887124413371	-0.246926308764	-0.149694810772	-0.524576567848	-0.518001420582	0.269697636269	0.217429939212	0.0831057120479	0.0226527785594	0.0889128003484	0.749728836895	0.134135336652	0.790663137616	-14.3061124043	5.41779163598	79.5179863007
10	2.30616582293	0.355053048245	-0.0287288943657	1.92375090535	0.222753471844	3.49934008307	0.121981128553	0.110232162665	0.211679889304	-0.160114244336	-0.18850798704	0.140880300218	0.0124249494197	0.139607194389	0.0368423226031	0.186367728801	0.0685944951785	0.0227245960919	0.493546552075	0.0501320060229	0.402499339204	-23.4158554061	-2.5137269928	79.2644131861
11	2.72847536944	1.77588393961	3.82907857192	2.30851526892	1.05202429656	7.17804587242	0.211523294974	-0.0449532452015	0.131469659111	-0.0375637672367	-0.536865683232	0.373262929831	-0.435778049247	-0.712328524097	0.303868822655	0.214472271422	0.101154916257	0.0223049416076	0.591503923241	0.0510177614274	0.499931818205	-28.3088001957	-0.857012091949	93.2311697031
12	4.31286698712	-0.840324823757	-0.278779615675	3.34413231414	0.917668922659	2.4494213036	0.00396073789706	0.494099343593	-0.20396642482	-0.351668529632	-0.220408151951	-0.605470322	0.694257050624	0.69227950139	0.0931080352604	0.363587124976	0.262317523292	0.0765569282274	0.706068538312	0.0583021852395	0.58160779772	-13.2743299173	-12.4623529283	75.0768327013
13	6.90769304993	-2.31404730277	-0.904069402111	8.3602041524	1.67743419744	3.6939806084	-0.128899563736	0.0442028078934	-1.1260134541	0.61420385809	-0.567883607818	-0.103160972035	1.73819171006	-0.237630861907	0.392139522238	0.716526172558	0.110858925289	-0.0252449265552	0.508876373624	0.0424761426158	0.523376405642	-22.1934239493	-20.4383053773	84.8627069321
14	3.796031859	0.959628235519	0.0304527213423	1.23932102046	0.355999011366	1.55221298375	0.233861501368	0.0537368909476	0.00816745839588	-0.135290752586	-0.272440586094	-0.548468261339	0.627077536769	0.549084346654	-0.0549049347722	0.250453116944	0.191255034828	0.0165245825418	0.530886294899	0.0574693047261	0.401012773008	-17.7143286526	-10.4538917759	80.8537597583
15	2.10902017194	0.535781889783	0.595446003281	2.54788086284	-0.344637147883	5.31664996292	0.0282448854034	0.0182878642813	0.26367153304	-0.349447832552	-0.0477185361909	0.133124990938	-0.560032487298	-0.507345092435	0.00152714441173	0.49174573324	0.247656504888	0.0286512855603	0.446262708927	0.05248364769	0.426882553614	-33.5274030177	2.27566620829	47.9825137351
16	1.87611551517	0.14974860054	0.293274070078	2.46540038172	-0.170280950232	5.50477541031	-0.0673475386406	-0.168918754852	-0.00632871733019	-0.244811575347	0.0676417974363	0.0185086856648	-0.0142600501942	-0.421447412755	-0.0229513774423	0.36582521323	0.172109065284	-0.018537589031	0.370695518004	0.0912329240898	0.362572090716	-33.7458179193	-3.93556810683	45.9288699082
17	2.74413628441	1.56719879511	1.77742855166	7.01020466456	-1.47224959071	2.17493840574	-0.316897694869	0.99908520328	0.663736704237	-0.957762428013	0.0631349537915	0.26349503287	-1.14940915878	-1.72368051126	0.220676852339	0.967693160654	0.170828363473	-0.0936567497806	0.912659018975	0.130104064322	0.65474473507	-43.1166281393	3.33590455195	8.91547542494
18	2.1347977313	0.956157049537	-0.0789172484746	1.74253501085	-1.15178703317	3.81035431318	-0.221401288156	0.389203856272	-0.223965907444	-0.990442579869	-0.0350261598487	-0.519426143115	1.5666670334	0.118476257708	0.142558860166	1.13789624088	0.107669536489	0.117310235502	0.498862984578	0.0496858044901	0.404571755077	-26.2698553129	-11.3598900176	6.70560542107

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	chain 'C' and (resid 101 through 117 )
2	X-RAY DIFFRACTION	2	chain 'C' and (resid 118 through 146 )
3	X-RAY DIFFRACTION	3	chain 'C' and (resid 147 through 192 )
4	X-RAY DIFFRACTION	4	chain 'C' and (resid 193 through 260 )
5	X-RAY DIFFRACTION	5	chain 'C' and (resid 261 through 284 )
6	X-RAY DIFFRACTION	6	chain 'A' and (resid 58 through 73 )
7	X-RAY DIFFRACTION	7	chain 'A' and (resid 74 through 100 )
8	X-RAY DIFFRACTION	8	chain 'A' and (resid 101 through 123 )
9	X-RAY DIFFRACTION	9	chain 'A' and (resid 124 through 146 )
10	X-RAY DIFFRACTION	10	chain 'A' and (resid 147 through 192 )
11	X-RAY DIFFRACTION	11	chain 'A' and (resid 193 through 206 )
12	X-RAY DIFFRACTION	12	chain 'A' and (resid 207 through 224 )
13	X-RAY DIFFRACTION	13	chain 'A' and (resid 225 through 249 )
14	X-RAY DIFFRACTION	14	chain 'A' and (resid 250 through 284 )
15	X-RAY DIFFRACTION	15	chain 'B' and (resid 58 through 146 )
16	X-RAY DIFFRACTION	16	chain 'B' and (resid 147 through 284 )
17	X-RAY DIFFRACTION	17	chain 'C' and (resid 58 through 73 )
18	X-RAY DIFFRACTION	18	chain 'C' and (resid 74 through 100 )

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