PDB-7k69: Crystal Structure of Dihydrofolate reductase (DHFR) from Mycobact...

ID or keywords:

Entry

Database: PDB / ID: 7k69

Title

Crystal Structure of Dihydrofolate reductase (DHFR) from Mycobacterium ulcerans Agy99 in complex with NADP and inhibitor SDDC-0001574

Components

Dihydrofolate reductase

Keywords

OXIDOREDUCTASE/OXIDOREDUCTASE INHIBITOR / SSGCID / SDDC / inhibitor / Structural Genomics / Seattle Structural Genomics Center for Infectious Disease / OXIDOREDUCTASE / OXIDOREDUCTASE-OXIDOREDUCTASE INHIBITOR complex

Function / homology

Function and homology information

dihydrofolate metabolic process / glycine biosynthetic process / dihydrofolate reductase / dihydrofolate reductase activity / folic acid metabolic process / tetrahydrofolate biosynthetic process / one-carbon metabolic process / NADP binding / cytosol
Similarity search - Function

Biological species

Mycobacterium ulcerans (bacteria)

Method

X-RAY DIFFRACTION /

MOLECULAR REPLACEMENT / Resolution: 1.6 Å

Authors

Seattle Structural Genomics Center for Infectious Disease (SSGCID)

Citation

Journal: To Be Published
Title: Crystal Structure of Dihydrofolate reductase (DHFR) from Mycobacterium ulcerans Agy99 in complex with NADP and inhibitor SDDC-0001574
Authors: Abendroth, J. / Dranow, D.M. / Santhakumar, V. / Walpole, C. / Lorimer, D.D. / Horanyi, P.S. / Edwards, T.E.

History

Deposition	Sep 19, 2020	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Mar 17, 2021	Provider: repository / Type: Initial release
Revision 1.1	Oct 18, 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

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	7k69.cif.gz	108.2 KB	Display	PDBx/mmCIF format
PDB format	pdb7k69.ent.gz	66.1 KB	Display	PDB format
PDBx/mmJSON format	7k69.json.gz		Tree view	PDBx/mmJSON format
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Validation report

Summary document	7k69_validation.pdf.gz	1.3 MB	Display	wwPDB validaton report
Full document	7k69_full_validation.pdf.gz	1.4 MB	Display
Data in XML	7k69_validation.xml.gz	11.6 KB	Display
Data in CIF	7k69_validation.cif.gz	16.9 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/k6/7k69 ftp://data.pdbj.org/pub/pdb/validation_reports/k6/7k69	HTTPS FTP

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Related structure data

Related structure data	6uwwS S: Starting model for refinement
Similar structure data	7k6a-d 5w3q-d 4e9b-d 5isp-d 3fy8-d 2w3v-d 5t6g-1 7lvc-d 3fyw-d 3f0q-d Search similar-shape structures by Omokage search (details) Similarity search - Function & homologyF&H Search Similarity search - FunctionF&H Search Similarity search - HomologyF&H Search
Other databases	TargetTrack: SSGCID-MyulA.01062.a.B12

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Links

PDB pages	PDBj / wwPDB / NCBI
Related items in Molecule of the Month	#34 - Oct 2002 Dihydrofolate Reductase similarity (8)

Deposited unit

A: Dihydrofolate reductase

hetero molecules

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

1

Idetical with deposited unit
defined by author&software
Evidence: gel filtration

Unit cell

Length a, b, c (Å)	28.710, 67.070, 44.110
Angle α, β, γ (deg.)	90.000, 91.337, 90.000
Int Tables number	4
Space group name H-M	P12₁1
Space group name Hall	P2yb
Symmetry operation	#1: x,y,z #2: -x,y+1/2,-z

#1: Protein	Dihydrofolate reductase Mass: 19051.684 Da / Num. of mol.: 1 / Mutation: E96A Source method: isolated from a genetically manipulated source Source: (gene. exp.) Mycobacterium ulcerans (strain Agy99) (bacteria) Strain: Agy99 / Gene: dfrA, MUL_2179 / Plasmid: MyulA.01062.a.B12 / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / References: UniProt: A0PQG8, dihydrofolate reductase
#2: Chemical	ChemComp-NAP / NADP NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE / 2'-MONOPHOSPHOADENOSINE 5'-DIPHOSPHORIBOSE Mass: 743.405 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₂₁H₂₈N₇O₁₇P₃ / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical	ChemComp-VY7 / 1-{2-[2-(2,4-diamino-5,7-dihydro-6H-pyrrolo[3,4-d]pyrimidin-6-yl)-2-oxoethoxy]phenyl}piperidine-4-carboxylic acid Mass: 412.442 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₂₀H₂₄N₆O₄ / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical	ChemComp-SER / SERINE Type: L-peptide linking / Mass: 105.093 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₃H₇NO₃
#5: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 218 / Source method: isolated from a natural source / Formula: H₂O
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: 2.23 Å³/Da / Density % sol: 44.9 %
Crystal grow	Temperature: 287 K / Method: vapor diffusion, sitting drop / pH: 6.5 Details: Morpheus screen, condition D2: 112.5% w/v PEG 1000, 12.5% w/v PEG 3350, 12.5% v/v MPD: 20mM of each sodium L-glutamate, DL-alanine, glycine, DL-lysine HCl, DL-serine: 100mM MES/imidazole pH ...Details: Morpheus screen, condition D2: 112.5% w/v PEG 1000, 12.5% w/v PEG 3350, 12.5% v/v MPD: 20mM of each sodium L-glutamate, DL-alanine, glycine, DL-lysine HCl, DL-serine: 100mM MES/imidazole pH 6.5: MyulA.01062.a.B12.PS38539 and 15.75mg/ml + 2.5mM NADP and 4mM SDDC compound SDDC-0001574: tray: 317895h4: cryo: direct: puck: xhf8-3

Crystal

Density Matthews: 2.23 Å³/Da / Density % sol: 44.9 %

Crystal grow

Temperature: 287 K / Method: vapor diffusion, sitting drop / pH: 6.5
Details: Morpheus screen, condition D2: 112.5% w/v PEG 1000, 12.5% w/v PEG 3350, 12.5% v/v MPD: 20mM of each sodium L-glutamate, DL-alanine, glycine, DL-lysine HCl, DL-serine: 100mM MES/imidazole pH ...

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

ROTATING ANODE / Type: RIGAKU FR-E+ SUPERBRIGHT / Wavelength: 1.5418 Å

Detector

Type: RIGAKU SATURN 944+ / Detector: CCD / Date: Sep 1, 2020

Radiation

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

Radiation wavelength

Wavelength: 1.5418 Å / Relative weight: 1

Reflection

Resolution: 1.6→50 Å / Num. obs: 20986 / % possible obs: 95 % / Redundancy: 6.326 % / B_iso Wilson estimate: 21.237 Å² / CC_1/2: 1 / R_merge(I) obs: 0.03 / R_rim(I) all: 0.033 / Χ²: 0.96 / Net I/σ(I): 35.22

Reflection shell

Diffraction-ID: 1

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Mean I/σ(I) obs	Num. unique obs	CC_1/2	R_rim(I) all	% possible all
1.6-1.64	4.205	0.225	6.44	1274	0.95	0.254	80
1.64-1.69	4.503	0.189	7.46	1380	0.972	0.212	86.2
1.69-1.74	4.626	0.158	8.76	1391	0.982	0.177	90.4
1.74-1.79	4.783	0.132	10.54	1345	0.989	0.148	88.8
1.79-1.85	4.865	0.103	13.5	1337	0.992	0.115	92.5
1.85-1.91	4.97	0.084	16.43	1341	0.996	0.094	93.4
1.91-1.98	5.003	0.067	20.57	1319	0.997	0.074	99.7
1.98-2.07	5.142	0.057	23.9	1278	0.998	0.063	97.3
2.07-2.16	5.377	0.048	29.16	1245	0.998	0.053	100
2.16-2.26	5.661	0.041	34.56	1198	0.999	0.045	99.8
2.26-2.39	6.519	0.04	38.71	1157	0.999	0.044	100
2.39-2.53	6.955	0.035	43.62	1069	0.999	0.038	100
2.53-2.7	7.322	0.031	51.28	1014	0.999	0.033	100
2.7-2.92	7.832	0.029	55.53	964	0.999	0.032	100
2.92-3.2	8.906	0.025	68.74	875	1	0.027	100
3.2-3.58	10.954	0.023	85.45	783	1	0.025	100
3.58-4.13	11.165	0.021	96.47	703	1	0.022	100
4.13-5.06	11.137	0.021	99.88	585	1	0.022	99.8
5.06-7.16	10.903	0.024	90.1	472	1	0.025	100
7.16-50	10.434	0.023	101.65	256	1	0.024	99.6

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Processing

Software

Name	Version	Classification
XDS		data reduction
XSCALE		data scaling
PHENIX	1.18.2	refinement
PDB_EXTRACT	3.25	data extraction
PHASER		phasing
Coot		model building

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: NAD and P218-bound structure, PDB entry 6uww

6uww

Resolution: 1.6→44.1 Å / SU ML: 0.1607 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 19.5376
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2

	R_factor	Num. reflection	% reflection	Selection details
R_free	0.1915	2117	10.09 %	0
R_work	0.1605	18866	-	-
obs	0.1637	20983	95.01 %	-

Solvent computation

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

Displacement parameters

B_iso mean: 18.22 Å²

Refinement step

Cycle: LAST / Resolution: 1.6→44.1 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	1232	0	85	218	1535

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.0086	1440
X-RAY DIFFRACTION	f_angle_d	1.1306	1996
X-RAY DIFFRACTION	f_chiral_restr	0.0585	218
X-RAY DIFFRACTION	f_plane_restr	0.0088	271
X-RAY DIFFRACTION	f_dihedral_angle_d	21.2928	554

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 (%)
1.6-1.64	0.2611	103	0.2171	1050	X-RAY DIFFRACTION	79.63
1.64-1.68	0.2209	128	0.1842	1164	X-RAY DIFFRACTION	86.48
1.68-1.72	0.2037	120	0.1669	1156	X-RAY DIFFRACTION	89.36
1.72-1.77	0.2454	121	0.1713	1204	X-RAY DIFFRACTION	89.47
1.77-1.83	0.215	134	0.168	1207	X-RAY DIFFRACTION	91.29
1.83-1.9	0.2024	157	0.1708	1186	X-RAY DIFFRACTION	93.07
1.9-1.97	0.2121	137	0.1676	1315	X-RAY DIFFRACTION	98.17
1.97-2.06	0.2048	139	0.1659	1301	X-RAY DIFFRACTION	97.56
2.06-2.17	0.1785	160	0.1644	1315	X-RAY DIFFRACTION	100
2.17-2.31	0.2058	164	0.1648	1293	X-RAY DIFFRACTION	99.86
2.31-2.49	0.2148	159	0.1747	1315	X-RAY DIFFRACTION	100
2.49-2.74	0.1723	148	0.1641	1329	X-RAY DIFFRACTION	100
2.74-3.13	0.2251	141	0.1633	1341	X-RAY DIFFRACTION	100
3.13-3.94	0.1768	149	0.1472	1330	X-RAY DIFFRACTION	100
3.95-44.1	0.1489	157	0.1383	1360	X-RAY DIFFRACTION	99.93

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	2.74563847789	0.172559235433	-0.204873596643	3.16310405423	0.187549246502	2.25275193237	-0.00498658194842	-0.201381568446	0.0709005738213	0.158118462944	-0.0142686963199	-0.14051504819	-0.211295438221	0.107243263609	0.0544454497364	0.0712998660614	-0.020452025985	-0.009324669053	0.0401848492294	-0.0128108922213	0.0936900216727	4.90260081177	0.923030217855	16.0966573518
2	2.20343473671	-0.826965741897	-0.141018562611	2.03894000478	1.08364708077	1.72328762086	-0.0805835337727	-0.275013502921	0.291775350134	-0.0544134115598	-0.182290890381	0.269772716176	-0.571580799874	-0.334631043555	0.0147646806316	0.12722830248	0.022009030297	-0.0201603949064	0.111047927622	-0.0418883296522	0.127946893406	-8.12104476632	1.89691109367	13.6713766646
3	2.87338187738	-1.15230952031	0.550094512861	2.47108581394	0.0696380763608	2.68543169447	-0.0479568713467	-0.185388583783	-0.118281514608	0.139118990999	0.0208072154275	0.0831182252482	0.0380796870666	-0.119298642159	0.044248087107	0.0600390535827	-0.0167910100118	0.0126326241019	0.0355906402523	0.0022674342606	0.0805232448298	-3.22730117364	-10.9564825476	13.0312895611
4	4.85107355993	-0.876487787295	-0.64221947687	3.07479097603	0.791776330038	2.02038603974	-0.100965388434	0.266421810907	-0.37509548061	-0.180444617086	-0.042575823926	-0.185805229271	0.250807312891	0.0957384842124	0.101648031385	0.141555289781	0.0135545274967	0.0312092362866	0.0797591865833	-0.0246789428665	0.144488855226	1.9922945797	-17.8980824555	6.08026993628
5	2.00612593487	-0.133101830995	-1.19602763788	5.56679220151	2.65137429719	2.55172544038	-0.0257554123188	0.477071958659	-0.269369628573	-0.954971805775	-0.220968949556	0.491176821228	-0.0143694640095	-0.52844864875	0.207923001186	0.217307477396	-0.0363454147089	-0.0660452132094	0.176084211142	-0.0292396836753	0.178239424757	-2.74129338663	-8.2324360655	0.283096348662
6	2.29798626341	-1.19225861875	-0.281474644224	2.54072165078	0.190169747538	1.77250642948	0.0488239116654	0.100860805954	0.0334472094339	-0.0787116224034	-0.142928044305	-0.112979658117	-0.175058724516	0.0803302940324	0.0710690635876	0.0790939601215	-0.0232019984359	-0.000318718707223	0.0493044601783	0.0159824311025	0.0890038443073	5.2571818768	0.976760154612	6.39612628442
7	6.06760606035	-4.99747195597	-1.97734354968	4.39227750348	1.92205834543	0.956118792089	-0.207922152328	-0.934069881491	0.855866278051	0.230734949594	0.672994301669	-1.23632381857	-0.144336653813	0.391688770745	-0.478441725355	0.174326594416	-0.00420036283611	-0.0514875579114	0.277666050031	-0.103360349898	0.288946406274	13.4382949892	1.50411780115	22.7451227248
8	0.0686728989173	-0.157843475115	0.522529631261	1.12966737804	-0.761091561475	4.16085310355	0.0512107790518	0.122184635238	0.245685839629	-0.053771750189	-0.172071916085	-0.290936093341	-0.549846395013	0.409615025948	0.0687411201839	0.164973356368	-0.0366586267238	0.0305138679152	0.127321876762	0.0401957887618	0.156295316412	8.52498471379	8.01929852031	3.5357786786
9	1.72585022663	-0.474459930442	-2.50660645661	0.91777550869	1.25449497856	5.50579624854	0.11907351575	0.0210752504766	0.133358612822	0.195180450651	-0.0861422729717	-0.139192391818	-0.254718857824	-0.196007223643	-0.0197626630973	0.26142387514	0.00948279000568	-0.00187102855939	0.0847277136244	-0.0172170427744	0.135482788477	-0.477508363989	11.6552338728	12.2603621039
10	2.00000063148	-5.75376752928	-5.00030673688	2.00000090838	1.99999877796	2.00000039468	1.18488105133	-0.401781056988	0.60479991709	5.04881704848	-1.34579706422	-0.672043516393	0.590218615965	0.674758906489	0.134098173267	0.643679487151	-0.0359428152688	-0.0588899396503	0.745158519487	0.0438597159685	0.623142410211	7.92180344859	10.6479800295	10.3239660058

Refinement TLS group

Refine-ID: X-RAY DIFFRACTION / Auth asym-ID: A

ID	Refine TLS-ID	Selection details	Label asym-ID	Auth seq-ID	Label seq-ID
1	1	chain 'A' and (resid 3 through 26 )	A	3 - 26	1 - 24
2	2	chain 'A' and (resid 27 through 37 )	A	27 - 37	25 - 35
3	3	chain 'A' and (resid 38 through 62 )	A	38 - 62	36 - 60
4	4	chain 'A' and (resid 63 through 87 )	A	63 - 87	61 - 85
5	5	chain 'A' and (resid 88 through 102 )	A	88 - 102	86 - 100
6	6	chain 'A' and (resid 103 through 122 )	A	103 - 122	101 - 120
7	7	chain 'A' and (resid 123 through 132 )	A	123 - 132	121 - 130
8	8	chain 'A' and (resid 133 through 145 )	A	133 - 145	131 - 143
9	9	chain 'A' and (resid 146 through 165 )	A	146 - 165	144 - 163
10	10	chain 'A' and (resid 301 through 301 )	C	301	2

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