PDB-5c5i: Crystal structure of NADP-dependent dehydrogenase from Rhodobacte...

ID or keywords:

Entry

Database: PDB / ID: 5c5i

Title

Crystal structure of NADP-dependent dehydrogenase from Rhodobacter sphaeroides

Components

NADP-dependent dehydrogenase

Keywords

OXIDOREDUCTASE / Structural Genomics / PSI-Biology / New York Structural Genomics Research Consortium / NYSGRC

Function / homology

Function and homology information

hydroxypyruvate reductase / glyoxylate reductase (NADP+) / hydroxypyruvate reductase [NAD(P)H] activity / glyoxylate reductase (NADPH) activity / NAD binding
Similarity search - Function

Biological species

Rhodobacter sphaeroides (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.2 Å

Authors

Kowiel, M. / Gasiorowska, O.A. / Shabalin, I.G. / Handing, K.B. / Porebski, P.J. / Cymborowski, M. / Al Obaidi, N.F. / Bonanno, J. / Almo, S.C. / Minor, W. / New York Structural Genomics Research Consortium (NYSGRC)

Funding support

United States, 1items

Citation

Journal: to be published
Title: Crystal structures of NADP-dependent dehydrogenase from Rhodobacter sphaeroides
Authors: Kowiel, M. / Gasiorowska, O.A. / Shabalin, I.G. / Handing, K.B. / Porebski, P.J. / Bonanno, J. / Almo, S.C. / Minor, W.

History

Deposition	Jun 19, 2015	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Jul 1, 2015	Provider: repository / Type: Initial release
Revision 1.1	Nov 22, 2017	Group: Database references / Derived calculations ...Database references / Derived calculations / Refinement description / Source and taxonomy Category: entity_src_gen / pdbx_database_related ...entity_src_gen / pdbx_database_related / pdbx_struct_assembly / pdbx_struct_assembly_gen / pdbx_struct_assembly_prop / pdbx_struct_oper_list / software Item: _entity_src_gen.pdbx_alt_source_flag / _pdbx_database_related.content_type ..._entity_src_gen.pdbx_alt_source_flag / _pdbx_database_related.content_type / _pdbx_struct_assembly.oligomeric_details / _pdbx_struct_assembly_gen.asym_id_list / _pdbx_struct_assembly_prop.type / _pdbx_struct_assembly_prop.value / _pdbx_struct_oper_list.symmetry_operation
Revision 1.2	Apr 13, 2022	Group: Database references / Structure summary / Category: audit_author / citation_author / database_2 Item: _audit_author.identifier_ORCID / _citation_author.identifier_ORCID ..._audit_author.identifier_ORCID / _citation_author.identifier_ORCID / _database_2.pdbx_DOI / _database_2.pdbx_database_accession
Revision 1.3	Sep 27, 2023	Group: Data collection / Refinement description Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model / struct_ncs_dom_lim Item: _struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id ..._struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id / _struct_ncs_dom_lim.beg_label_comp_id / _struct_ncs_dom_lim.beg_label_seq_id / _struct_ncs_dom_lim.end_auth_comp_id / _struct_ncs_dom_lim.end_label_asym_id / _struct_ncs_dom_lim.end_label_comp_id / _struct_ncs_dom_lim.end_label_seq_id

Structure viewer	Molecule: MolmilJmol/JSmol

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Validation report

Summary document	5c5i_validation.pdf.gz	429.6 KB	Display	wwPDB validaton report
Full document	5c5i_full_validation.pdf.gz	432.9 KB	Display
Data in XML	5c5i_validation.xml.gz	22.8 KB	Display
Data in CIF	5c5i_validation.cif.gz	32.2 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/c5/5c5i ftp://data.pdbj.org/pub/pdb/validation_reports/c5/5c5i	HTTPS FTP

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Related structure data

Related structure data	4zqbS S: Starting model for refinement
Similar structure data	6brg-3 5mh6-1 5mha-d 6hon-d 6hom-d 6brh-1 5mh6-2 6rih-d 6brh-3 4xa8-1 Search similar-shape structures by Omokage search (details) Similarity search - Function & homologyF&H Search Similarity search - FunctionF&H Search Similarity search - HomologyF&H Search
Other databases	TargetTrack: NYSGRC-014667

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Links

PDB pages	PDBj / wwPDB / NCBI

Deposited unit

A: NADP-dependent dehydrogenase

B: NADP-dependent dehydrogenase

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

1

Idetical with deposited unit
defined by author&software

Unit cell

Length a, b, c (Å)	110.048, 80.085, 61.330
Angle α, β, γ (deg.)	90.000, 93.690, 90.000
Int Tables number	5
Space group name H-M	C121

Noncrystallographic symmetry (NCS)

NCS domain:

ID	Ens-ID	Details
1	1	A
2	1	B

NCS domain segments:

Component-ID: _ / Ens-ID: 1 / Beg auth comp-ID: SER / Beg label comp-ID: SER / End auth comp-ID: VAL / End label comp-ID: VAL / Refine code: _ / Auth seq-ID: 0 - 307 / Label seq-ID: 3 - 310

Dom-ID	Auth asym-ID	Label asym-ID
1	A	A
2	B	B

#1: Protein	NADP-dependent dehydrogenase Mass: 34282.211 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Details: Before crystallization protein was incubated with 1/50 v/v of 2 mg/ml chymotrypsin. Source: (gene. exp.) Rhodobacter sphaeroides (bacteria) / Strain: ATCC 17023 / 2.4.1 / NCIB 8253 / DSM 158 / Gene: RSP_3442 / Plasmid: pSGC-His / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 (DE3) RIL References: UniProt: Q3IWN8, glyoxylate reductase (NADP+), hydroxypyruvate reductase
#2: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 103 / Source method: isolated from a natural source / Formula: H₂O

#1: Protein

NADP-dependent dehydrogenase

Mass: 34282.211 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: Before crystallization protein was incubated with 1/50 v/v of 2 mg/ml chymotrypsin.
Source: (gene. exp.)

Rhodobacter sphaeroides (bacteria) / Strain: ATCC 17023 / 2.4.1 / NCIB 8253 / DSM 158 / Gene: RSP_3442 / Plasmid: pSGC-His / Production host:

Escherichia coli (E. coli) / Strain (production host): BL21 (DE3) RIL
References: UniProt: Q3IWN8, glyoxylate reductase (NADP+), hydroxypyruvate reductase

#2: Water

ChemComp-HOH / water

Mass: 18.015 Da / Num. of mol.: 103 / Source method: isolated from a natural source / Formula: H₂O

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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 Å³/Da / Density % sol: 38.54 %
Crystal grow	Temperature: 289 K / Method: vapor diffusion, sitting drop / pH: 7.5 Details: 0.2 ul of 10.5 mg/ml protein in 20 mM HEPES pH 7.5, 150 mM NaCl, 10% Glycerol, 0.1% Sodium Azide and 0.5 mM TCEP were mixed with 0.2 ul of the MCSG Suite 2 condition # 72 (20%w/v PEG 3350, 0. ...Details: 0.2 ul of 10.5 mg/ml protein in 20 mM HEPES pH 7.5, 150 mM NaCl, 10% Glycerol, 0.1% Sodium Azide and 0.5 mM TCEP were mixed with 0.2 ul of the MCSG Suite 2 condition # 72 (20%w/v PEG 3350, 0.2M Li nitrate) and equilibrated against 1.5 M NaCl solution in 96 Well 3 drop Crystallization Plate (Swissci). Before crystallization protein was incubated with 1/50 v/v of 2 mg/ml chymotrypsin

Crystal

Density Matthews: 2 Å³/Da / Density % sol: 38.54 %

Crystal grow

Temperature: 289 K / Method: vapor diffusion, sitting drop / pH: 7.5
Details: 0.2 ul of 10.5 mg/ml protein in 20 mM HEPES pH 7.5, 150 mM NaCl, 10% Glycerol, 0.1% Sodium Azide and 0.5 mM TCEP were mixed with 0.2 ul of the MCSG Suite 2 condition # 72 (20%w/v PEG 3350, 0. ...

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Data collection

Diffraction

Mean temperature: 100 K

Diffraction source

Source:

SYNCHROTRON / Site:

APS

/ Beamline: 21-ID-G / Wavelength: 0.97856 Å

Detector

Type: MARMOSAIC 300 mm CCD / Detector: CCD / Date: Nov 6, 2014

Radiation

Monochromator: Diamond [111] / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray

Radiation wavelength

Wavelength: 0.97856 Å / Relative weight: 1

Reflection

Resolution: 2.2→50 Å / Num. obs: 26059 / % possible obs: 96.4 % / Observed criterion σ(I): -3 / Redundancy: 4.4 % / B_iso Wilson estimate: 41.3 Å² / R_merge(I) obs: 0.074 / R_pim(I) all: 0.04 / R_rim(I) all: 0.085 / R_sym value: 0.074 / Χ²: 1.499 / Net I/av σ(I): 25.582 / Net I/σ(I): 8.7 / Num. measured all: 113742

Reflection shell

Diffraction-ID: 1 / Rejects: _

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Mean I/σ(I) obs	Num. unique all	CC_1/2	R_pim(I) all	R_rim(I) all	Χ²	% possible all
2.2-2.24	3.9	0.527	2.2	1079	0.893	0.286	0.602	0.76	79.5
2.24-2.28	4	0.485		1137	0.905	0.257	0.551	0.852	84.8
2.28-2.32	4.1	0.433		1154	0.934	0.229	0.492	0.841	87.7
2.32-2.37	4.2	0.383		1245	0.947	0.203	0.435	0.828	92.2
2.37-2.42	4.3	0.359		1298	0.971	0.189	0.407	0.884	96.2
2.42-2.48	4.4	0.349		1343	0.96	0.18	0.394	0.935	99.4
2.48-2.54	4.5	0.303		1351	0.97	0.157	0.343	0.942	99.9
2.54-2.61	4.6	0.258		1343	0.976	0.133	0.291	1.031	100
2.61-2.69	4.7	0.218		1327	0.987	0.113	0.246	1.083	99.9
2.69-2.77	4.7	0.184		1352	0.986	0.095	0.207	1.225	100
2.77-2.87	4.7	0.148		1359	0.991	0.076	0.167	1.316	99.7
2.87-2.99	4.6	0.132		1340	0.992	0.068	0.149	1.452	100
2.99-3.12	4.6	0.107		1343	0.994	0.055	0.121	1.561	99.9
3.12-3.29	4.5	0.083		1334	0.996	0.044	0.095	1.675	99.8
3.29-3.49	4.4	0.072		1357	0.995	0.039	0.082	1.952	98.8
3.49-3.76	4.3	0.067		1337	0.994	0.037	0.077	2.415	98.6
3.76-4.14	4.2	0.057		1299	0.995	0.032	0.066	2.544	96.7
4.14-4.74	4	0.054		1330	0.996	0.031	0.063	2.812	96.9
4.74-5.97	4.3	0.047		1354	0.997	0.025	0.054	2.337	99.1
5.97-50	4.1	0.042		1377	0.996	0.023	0.048	2.339	98.2

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Processing

Software

Name	Version	Classification
BLU-MAX		data collection
HKL-3000		data reduction
HKL-3000		phasing
HKL-3000		data scaling
MOLREP		phasing
REFMAC	5.8.0107	refinement
PDB_EXTRACT	3.15	data extraction

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 4ZQB

4zqb

Resolution: 2.2→50 Å / Cor.coef. F_o:F_c: 0.964 / Cor.coef. F_o:F_c free: 0.938 / WR_factor R_free: 0.2498 / WR_factor R_work: 0.1866 / FOM work R set: 0.7138 / SU B: 24.162 / SU ML: 0.266 / SU R Cruickshank DPI: 0.3836 / SU R_free: 0.2475 / Cross valid method: THROUGHOUT / σ(F): 0 / ESU R: 0.384 / ESU R Free: 0.248 / Stereochemistry target values: MAXIMUM LIKELIHOOD
Details: U VALUES : WITH TLS ADDED HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS

	R_factor	Num. reflection	% reflection	Selection details
R_free	0.252	1253	4.8 %	RANDOM
R_work	0.1945	-	-	-
obs	0.1973	26059	96.36 %	-

Solvent computation

Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK

Displacement parameters

B_iso max: 126.55 Å² / B_iso mean: 62.532 Å² / B_iso min: 17.82 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	-1.11 Å²	0 Å²	1.16 Å²
2-	-	7.09 Å²	-0 Å²
3-	-	-	-6.08 Å²

Refinement step

Cycle: final / Resolution: 2.2→50 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4537	0	0	104	4641
B_iso mean	-	-	-	54.98	-
Num. residues	-	-	-	-	620

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.014	0.019	4681
X-RAY DIFFRACTION	r_bond_other_d	0.006	0.02	4451
X-RAY DIFFRACTION	r_angle_refined_deg	1.657	1.965	6431
X-RAY DIFFRACTION	r_angle_other_deg	1.271	3	10195
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	5.807	5	626
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	36.004	22.857	168
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	13.702	15	653
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	22.018	15	31
X-RAY DIFFRACTION	r_chiral_restr	0.089	0.2	755
X-RAY DIFFRACTION	r_gen_planes_refined	0.009	0.021	5332
X-RAY DIFFRACTION	r_gen_planes_other	0.005	0.02	1003
X-RAY DIFFRACTION	r_mcbond_it	1.579	3.538	2486
X-RAY DIFFRACTION	r_mcbond_other	1.58	3.538	2485
X-RAY DIFFRACTION	r_mcangle_it	2.457	5.304	3106

Refine LS restraints NCS

Ens-ID: 1 / Number: 33128 / Refine-ID: X-RAY DIFFRACTION / Type: interatomic distance / Rms dev position: 0.11 Å / Weight position: 0.05

Dom-ID	Auth asym-ID
1	A
2	B

LS refinement shell

Resolution: 2.201→2.258 Å / Total num. of bins used: 20

	R_factor	Num. reflection	% reflection
R_free	0.368	78	-
R_work	0.343	1511	-
all	-	1589	-
obs	-	-	80.17 %

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	12.8918	-7.3754	-2.026	6.345	6.5164	13.9761	0.8333	-0.5488	1.8469	-0.2996	-0.3207	-1.2208	0.204	-1.3207	-0.5127	0.3664	-0.1579	-0.0031	0.5934	0.1313	0.5778	9.783	23.204	56.827
2	0.0384	0.205	0.2066	3.3381	3.7499	4.3676	0.0929	0.0209	-0.0626	0.1741	0.1597	-0.3959	0.2027	0.0518	-0.2525	0.2774	0.0083	-0.0094	0.1714	-0.0617	0.396	20.067	12.78	39.996
3	2.1116	0.7167	-0.4319	7.7823	-0.9173	4.0325	0.0582	-0.3107	0.0192	0.8417	-0.1559	-0.2247	0.2424	-0.1981	0.0977	0.2976	-0.0399	0.0249	0.2055	0.0507	0.0563	11.764	12.026	46.997
4	13.527	14.1798	-0.0765	14.9658	-0.1227	0.0525	-0.2658	0.1177	-1.1124	-0.4846	0.2715	-1.0052	0.0712	-0.1933	-0.0057	0.7353	-0.1178	-0.1361	0.7634	0.0917	0.5576	3.526	2.827	39.582
5	0.7987	0.0429	0.0776	0.1078	-0.3554	2.3323	0.0955	0.2447	-0.0335	0.0223	-0.0548	0.0049	-0.0022	-0.0215	-0.0407	0.305	-0.0998	0.0035	0.2274	-0.0316	0.2545	23.374	1.812	15.35
6	4.6822	0.5685	-0.8945	1.9254	-0.5847	4.7266	0.0502	0.2119	-0.0693	-0.2125	0.029	0.2667	-0.1585	-0.5587	-0.0791	0.2107	-0.067	-0.0681	0.2636	0.0218	0.2061	9.523	3.298	6.057
7	3.4236	-1.4451	0.1617	2.0678	-1.0775	3.527	-0.0949	0.3601	-0.1941	-0.0161	0.127	0.5733	0.5788	-0.6457	-0.0321	0.2372	-0.1735	0.0048	0.2544	-0.0019	0.2613	7.815	-7.739	12.485
8	1.851	0.2967	-0.4574	2.7154	-0.0213	3.7969	-0.0832	-0.0943	-0.1476	-0.1289	0.0141	0.3665	0.369	-0.1471	0.0692	0.2871	-0.0532	-0.0166	0.0838	0.0053	0.2071	17.823	-8.564	23.666
9	2.4848	0.0099	0.2678	3.4135	1.7712	4.3979	0.214	-0.0921	0.2288	-0.381	-0.0479	0.4984	-0.2721	-1.0203	-0.1661	0.2186	0.0663	-0.0005	0.3101	0.0856	0.2364	7.273	19.562	36.063
10	1.897	2.5075	-0.4514	3.7768	-0.6116	10.5628	0.2267	-0.0335	0.0284	0.1776	0.2605	-0.0865	0.8657	-0.3098	-0.4873	0.3009	0.106	-0.0243	0.5135	-0.0309	0.1608	40.962	-11.37	-9.863
11	7.7988	-2.3346	1.2429	2.3351	-1.2413	4.685	0.0973	0.1637	-0.5363	-0.2687	0.0425	0.0815	0.24	0.2064	-0.1397	0.2948	0.0604	-0.0135	0.3041	-0.0198	0.1464	35.207	-10.727	-13.385
12	5.6433	4.0396	1.9253	27.8567	-1.7234	4.1892	0.0724	-0.2191	-0.5894	-0.2614	-0.0461	-1.01	0.3332	0.2008	-0.0264	0.2581	0.3	0.1135	0.5459	0.0573	0.2767	47.182	-16.611	-16.897
13	3.694	0.5763	-0.0489	3.921	-1.6978	0.7494	-0.0443	-0.6707	0.227	0.0879	-0.1521	-0.4392	-0.0268	0.0735	0.1965	0.1684	0.1821	-0.09	0.7846	-0.1173	0.1729	46.811	-4.598	-11.295
14	12.6909	-4.706	-7.2956	2.2458	1.9324	5.5111	0.3451	-0.1506	0.3579	-0.0908	-0.146	-0.2119	-0.068	0.3621	-0.1991	0.3063	-0.0723	-0.1375	0.3336	-0.0806	0.3366	43.429	4.468	-8.272
15	0.9566	0.0142	-0.8949	2.2203	-0.3716	2.9513	-0.0042	-0.2817	0.0068	0.1751	-0.0182	-0.0983	-0.1958	0.2945	0.0223	0.2683	-0.074	-0.0254	0.143	-0.0152	0.2139	27.309	4.149	21.901
16	2.3416	-0.5884	-0.3113	3.7827	-1.7607	1.2255	-0.0017	0.252	0.1406	0.0609	-0.1448	-0.3016	-0.3039	0.2977	0.1466	0.353	-0.3088	0.0125	0.3625	0.0459	0.1798	34.797	20.488	9.135
17	5.1038	1.1997	1.2206	2.2184	-0.3932	3.4827	-0.0433	0.4447	-0.0053	0.2491	0.0252	-0.6151	-0.2261	0.6186	0.0181	0.2115	-0.2479	-0.0548	0.3947	0.0055	0.3763	44.398	13.699	14.598
18	3.545	-1.9617	0.597	2.8524	0.324	0.3446	-0.094	-0.1081	0.5022	0.0758	0.0725	-0.4338	-0.0094	0.0028	0.0214	0.0896	-0.1506	-0.0237	0.4399	0.0506	0.221	45.097	6.253	17.369
19	2.5391	-0.3045	2.041	1.1865	-0.1902	10.2529	0.0265	0.1248	-0.1127	-0.0697	-0.1405	-0.1018	-0.0056	0.2883	0.1141	0.1506	-0.0548	0.0191	0.2064	0.0009	0.2183	40.128	-1.766	8.765
20	4.7106	-0.0985	-2.5308	4.0072	-0.6495	2.1793	0.04	-0.1812	0.6368	-0.0817	0.0533	-0.0431	-0.1827	0.1792	-0.0934	0.2523	0.0075	-0.0866	0.304	-0.0288	0.1795	39.368	1.609	-20.669

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	A	-2 - 3
2	X-RAY DIFFRACTION	2	A	4 - 25
3	X-RAY DIFFRACTION	3	A	26 - 71
4	X-RAY DIFFRACTION	4	A	72 - 85
5	X-RAY DIFFRACTION	5	A	86 - 133
6	X-RAY DIFFRACTION	6	A	134 - 178
7	X-RAY DIFFRACTION	7	A	179 - 227
8	X-RAY DIFFRACTION	8	A	228 - 280
9	X-RAY DIFFRACTION	9	A	281 - 308
10	X-RAY DIFFRACTION	10	B	0 - 15
11	X-RAY DIFFRACTION	11	B	16 - 35
12	X-RAY DIFFRACTION	12	B	36 - 46
13	X-RAY DIFFRACTION	13	B	47 - 74
14	X-RAY DIFFRACTION	14	B	75 - 101
15	X-RAY DIFFRACTION	15	B	102 - 133
16	X-RAY DIFFRACTION	16	B	134 - 205
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