PDB-7k6a: Crystal Structure of Dihydrofolate reductase (DHFR) from Mycobact...

ID or keywords:

Entry

Database: PDB / ID: 7k6a

Title

Crystal Structure of Dihydrofolate reductase (DHFR) from Mycobacterium ulcerans Agy99 in complex with NADP and inhibitor SDDC-0001575

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-0001575
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 ...Data collection / Database references / Refinement description / Structure summary Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_keywords Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_keywords.text

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	7k6a.cif.gz	108.3 KB	Display	PDBx/mmCIF format
PDB format	pdb7k6a.ent.gz	65.6 KB	Display	PDB format
PDBx/mmJSON format	7k6a.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	7k6a_validation.pdf.gz	1.3 MB	Display	wwPDB validaton report
Full document	7k6a_full_validation.pdf.gz	1.3 MB	Display
Data in XML	7k6a_validation.xml.gz	11.3 KB	Display
Data in CIF	7k6a_validation.cif.gz	16.6 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/k6/7k6a ftp://data.pdbj.org/pub/pdb/validation_reports/k6/7k6a	HTTPS FTP

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Related structure data

Related structure data	6uwwS S: Starting model for refinement
Similar structure data	7k69-d 7k68-d 6vv7-2 6nni-2 2w3v-d 7lvc-d 5w3q-d 3sru-d 3fqf-d 5z6l-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	TargetDB: 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

20.5 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)	28.870, 66.650, 44.290
Angle α, β, γ (deg.)	90.000, 91.800, 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

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Protein , 1 types, 1 molecules A

#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

#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

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Non-polymers , 5 types, 211 molecules

#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
#3: Chemical	ChemComp-VYG / 6-ethyl-5-[3-({3-[4-(trifluoromethyl)piperidin-1-yl]pyridin-2-yl}oxy)propoxy]pyrimidine-2,4-diamine Mass: 440.463 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₂₀H₂₇F₃N₆O₂ / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical	ChemComp-CL / CHLORIDE ION Mass: 35.453 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Cl / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical	ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL Mass: 62.068 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C₂H₆O₂
#6: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 205 / 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: 2.23 Å³/Da / Density % sol: 44.9 %
Crystal grow	Temperature: 287 K / Method: vapor diffusion, sitting drop / pH: 6.5 Details: Morpheus screen, condition F2: 12.5% w/v PEG 1000, 12.5% w/v PEG 3350, 12.5% v/v MPD: 20mM of each D-glucose, D-mannose, D-galactose, L-fucose, D-xylose, N-acetyl-D-glucosamine: 100mM ...Details: Morpheus screen, condition F2: 12.5% w/v PEG 1000, 12.5% w/v PEG 3350, 12.5% v/v MPD: 20mM of each D-glucose, D-mannose, D-galactose, L-fucose, D-xylose, N-acetyl-D-glucosamine: 100mM MES/imidazole pH 6.5: MyulA.01062.a.B12.PS38539 and 15.75mg/ml + 2.5mM NADP and 4mM SDDC compound SDDC-0001575: tray: 317893F2: cryo: direct: puck: xhf8-7

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 F2: 12.5% w/v PEG 1000, 12.5% w/v PEG 3350, 12.5% v/v MPD: 20mM of each D-glucose, D-mannose, D-galactose, L-fucose, D-xylose, N-acetyl-D-glucosamine: 100mM ...

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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 2, 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: 21861 / % possible obs: 98.6 % / Redundancy: 6.162 % / B_iso Wilson estimate: 19.18 Å² / CC_1/2: 0.999 / R_merge(I) obs: 0.047 / R_rim(I) all: 0.051 / Χ²: 0.929 / Net I/σ(I): 25.21

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	3.696	0.4	3.62	1524	0.879	0.464	94.6
1.64-1.69	4.206	0.329	4.45	1522	0.916	0.376	94.4
1.69-1.74	4.346	0.301	5.18	1476	0.928	0.341	96.2
1.74-1.79	4.401	0.239	6.43	1481	0.959	0.271	98
1.79-1.85	4.599	0.194	8.1	1445	0.974	0.219	98.8
1.85-1.91	4.743	0.158	9.67	1400	0.983	0.178	98.9
1.91-1.98	4.975	0.115	13.42	1334	0.991	0.128	99.9
1.98-2.07	5.088	0.1	16.2	1334	0.993	0.111	100
2.07-2.16	5.371	0.084	19.29	1230	0.996	0.093	100
2.16-2.26	5.817	0.07	24.06	1218	0.997	0.077	100
2.26-2.39	6.628	0.065	27.26	1156	0.998	0.071	99.9
2.39-2.53	7.019	0.057	31	1065	0.998	0.062	100
2.53-2.7	7.387	0.051	35.89	1017	0.998	0.055	99.9
2.7-2.92	8.012	0.045	40.35	973	0.999	0.048	100
2.92-3.2	9.039	0.036	52.69	869	0.999	0.038	100
3.2-3.58	11.076	0.03	69.86	791	1	0.031	100
3.58-4.13	11.134	0.024	83.77	707	1	0.025	100
4.13-5.06	11.138	0.024	84.13	589	1	0.025	100
5.06-7.16	11.011	0.032	66.87	470	1	0.034	100
7.16-50	10.292	0.027	86.34	260	1	0.028	100

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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→44.27 Å / SU ML: 0.1368 / Cross valid method: FREE R-VALUE / σ(F): 1.35 / Phase error: 17.4672
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2

	R_factor	Num. reflection	% reflection	Selection details
R_free	0.1784	2074	9.49 %	0
R_work	0.1464	19781	-	-
obs	0.1494	21855	98.63 %	-

Solvent computation

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

Displacement parameters

B_iso mean: 15.55 Å²

Refinement step

Cycle: LAST / Resolution: 1.6→44.27 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	1232	0	92	205	1529

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.0072	1425
X-RAY DIFFRACTION	f_angle_d	1.084	1969
X-RAY DIFFRACTION	f_chiral_restr	0.0581	217
X-RAY DIFFRACTION	f_plane_restr	0.01	268
X-RAY DIFFRACTION	f_dihedral_angle_d	13.5904	546

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	0.2551	122	0.2053	1252	X-RAY DIFFRACTION	94.63
1.64-1.68	0.2208	135	0.1798	1274	X-RAY DIFFRACTION	94.95
1.68-1.72	0.219	117	0.1676	1279	X-RAY DIFFRACTION	95.16
1.72-1.77	0.1968	144	0.1593	1299	X-RAY DIFFRACTION	98.03
1.77-1.83	0.1924	113	0.1506	1311	X-RAY DIFFRACTION	98
1.83-1.9	0.1654	141	0.149	1316	X-RAY DIFFRACTION	98.98
1.9-1.97	0.1957	172	0.142	1299	X-RAY DIFFRACTION	99.66
1.97-2.06	0.1537	124	0.1373	1386	X-RAY DIFFRACTION	100
2.06-2.17	0.1531	126	0.1389	1321	X-RAY DIFFRACTION	100
2.17-2.31	0.166	148	0.1443	1319	X-RAY DIFFRACTION	100
2.31-2.49	0.1954	150	0.1429	1337	X-RAY DIFFRACTION	99.93
2.49-2.74	0.1771	142	0.1459	1334	X-RAY DIFFRACTION	100
2.74-3.13	0.1724	157	0.147	1328	X-RAY DIFFRACTION	100
3.13-3.94	0.1662	159	0.1321	1331	X-RAY DIFFRACTION	100
3.95-44.27	0.1712	124	0.1449	1395	X-RAY DIFFRACTION	100

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	2.78055404451	-0.0478547672929	0.195710165339	2.92595795099	-0.203185351852	3.34381322031	0.0573633615491	-0.206339687659	0.0958057114998	0.103676182507	-0.0267127704155	-0.100712525234	-0.0692665103194	-0.00977462906644	-0.0153503921658	0.0459573006089	-0.00569581404069	0.00804476761752	0.0438007601257	-0.0059470820304	0.065783592315	5.18804965196	1.81815805988	16.1283077721
2	3.29491929859	-0.523957495708	-0.0122643639328	4.42450950718	3.2798274404	4.63522873144	0.0728970396705	-0.163082921276	0.187797253822	-0.0927501783669	-0.185743100918	0.232685180196	-0.245321628355	-0.0649907615034	0.0894311412195	0.0595563990217	0.00303836169961	0.00100435089926	0.0509493538441	-0.0122474429597	0.0885009468837	-7.91341795665	2.87319239703	13.5374747624
3	4.06838310235	-1.25712664848	0.486362241409	2.693348463	-0.694080446125	3.0054534564	-0.0151098364777	-0.0839921534131	-0.0657984932181	0.0434013604897	-0.0267399009161	0.0986303040908	0.0428076624658	-0.0395018002896	0.0399056683397	0.0541714212324	-0.0117119248612	0.00537321179929	0.0199128022747	-0.0168214979919	0.052152124532	-3.28636423202	-10.1335724292	12.8667689008
4	4.33454446246	-1.3857548596	-0.721360234631	3.7245940039	-0.266591965578	1.51023449656	-0.207089563726	0.287916917257	-0.411341673083	0.0156381041339	-0.00739606916867	-0.245475670161	0.254167706112	-0.0617205176445	0.111873424203	0.120627583795	0.00975593568885	0.0275425485292	0.0527401712075	-0.0338536644185	0.13353661025	0.988785139739	-18.3742976266	8.80868348037
5	2.43391745978	0.314745734884	-0.593965892568	7.19227979314	1.12268482792	3.97515508863	0.00269775105948	0.149209587714	0.280897676756	-0.537172534459	0.123005872017	-0.443948765599	-0.0125589751452	0.313149776242	0.000594477460274	0.136260418696	0.0063203208402	0.0190706905284	0.124034464295	-0.027133546483	0.111910102676	3.40942475309	-15.1917356577	1.59061014083
6	0.576330751105	0.402521298341	-0.636615382091	6.19250811113	2.52275180297	3.03126021068	-0.234814741985	0.266554725309	-0.292137847333	-0.922000976114	-0.028472897806	0.494011107336	-0.261565048049	-0.306944090126	0.0686550900401	0.197612911948	-0.0225496521396	-0.0502811880062	0.134384951853	-0.0356211049647	0.165710238584	-2.57098970664	-7.54415306298	0.350396803164
7	3.39013868856	-1.2835011726	1.17183773597	3.69405563166	-2.73251475958	5.90041004735	0.144495118543	0.331699874531	-0.0127925165075	-0.303028558551	-0.375843698747	-0.0486187626303	0.0572655729003	0.294877648845	0.19505479522	0.0741693343485	0.0226888097518	0.0135353532755	0.0841095378924	0.00423715295994	0.0928627837534	9.04807105685	-4.1773441871	4.16769559766
8	1.94190610756	-1.58724512553	-0.88609339014	2.70799830685	0.237563497524	0.389105474855	-0.182117098414	-0.183728345021	0.237805128437	0.158159585819	0.0954768064665	-0.311456845129	-0.131029000863	-0.00921449137632	0.0547537129542	0.0784637626352	-0.0209210357584	-0.0238232558584	0.0440762922029	-0.00688898729051	0.112785804343	7.21057018878	4.70887078303	14.6912206285
9	0.138418876128	-0.234893943974	0.683181724594	1.01686104181	-0.365513868294	3.15422322219	0.101441795236	0.222705484063	0.126998936055	-0.0481823875531	-0.193552352998	-0.143055937641	-0.508988872991	0.418810476781	0.0686284576523	0.139084284879	-4.56840427273E-5	0.0357852237703	0.119533045428	0.0468796036764	0.124906575047	8.5811147371	8.72700319898	3.46077632865
10	2.04980492328	-0.51395406028	-3.06129686413	1.62967892122	2.51743084244	7.21846556132	0.107371699389	0.0888744433362	0.0953395430677	-0.0878477680395	-0.0924684712434	-0.00696890977417	-0.100525478017	-0.203800584912	-0.013566493221	0.123461398173	0.0152393220142	0.000505051909494	0.0503977601201	0.00456780548997	0.091608954004	-0.101581269116	11.8661584442	10.9204890595

Refinement TLS group

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

ID	Refine TLS-ID	Selection details	Auth seq-ID	Label seq-ID
1	1	chain 'A' and (resid 3 through 26 )	3 - 26	1 - 24
2	2	chain 'A' and (resid 27 through 37 )	27 - 37	25 - 35
3	3	chain 'A' and (resid 38 through 62 )	38 - 62	36 - 60
4	4	chain 'A' and (resid 63 through 78 )	63 - 78	61 - 76
5	5	chain 'A' and (resid 79 through 87 )	79 - 87	77 - 85
6	6	chain 'A' and (resid 88 through 102 )	88 - 102	86 - 100
7	7	chain 'A' and (resid 103 through 112 )	103 - 112	101 - 110
8	8	chain 'A' and (resid 113 through 132 )	113 - 132	111 - 130
9	9	chain 'A' and (resid 133 through 145 )	133 - 145	131 - 143
10	10	chain 'A' and (resid 146 through 165 )	146 - 165	144 - 163

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Protein , 1 types, 1 molecules A

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Non-polymers , 5 types, 211 molecules

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Details

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Experimental details

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Experiment

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Sample preparation

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Data collection

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Processing

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