PDB-9qwy: Crystal structure of an NtA622L variant in complex with NADPH and...

ID or keywords:

Entry

Database: PDB / ID: 9qwy

Title

Crystal structure of an NtA622L variant in complex with NADPH and Trigonelline

Components

Isoflavone reductase homolog A622-like

Keywords

PLANT PROTEIN / Oxidoreductase / Nicotine / Niacin / NADPH / Rossmann-fold / IFR-like / PIP-reductase

Function / homology

Function and homology information

nicotine biosynthetic process / lignan biosynthetic process / oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor / Oxidoreductases; Acting on the CH-CH group of donors; With NAD+ or NADP+ as acceptor / alkaloid metabolic process / secondary metabolite biosynthetic process / cytoplasm
Similarity search - Function

Biological species

Nicotiana tabacum (common tobacco)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.6 Å

Authors

Mokos, D. / Leitner, D. / Daniel, B.

Funding support

Austria, 2items

Citation

Journal: To Be Published
Title: A622: a key oxidoreductase in the nicotine biosynthesis pathway
Authors: Mokos, D. / Leitner, D. / Schruefer, A. / Grubmueller, S. / Reisz, C. / Juric, J. / Berger, T. / Singh, A. / Friess, M. / Breinbauer, R. / Gruber, K. / Daniel, B.

History

Deposition	Apr 15, 2025	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Apr 29, 2026	Provider: repository / Type: Initial release

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	9qwy.cif.gz	873.5 KB	Display	PDBx/mmCIF format
PDB format	pdb9qwy.ent.gz		Display	PDB format
PDBx/mmJSON format	9qwy.json.gz		Tree view	PDBx/mmJSON format
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Validation report

Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/qw/9qwy ftp://data.pdbj.org/pub/pdb/validation_reports/qw/9qwy	HTTPS FTP

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Related structure data

Related structure data	9qrg 9qsw
Similar structure data	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	#34 - Oct 2002 Dihydrofolate Reductase similarity (1) #57 - Sep 2004 Catalase similarity (1)

Deposited unit

A: Isoflavone reductase homolog A622-like

B: Isoflavone reductase homolog A622-like

C: Isoflavone reductase homolog A622-like

hetero molecules

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

1

A: Isoflavone reductase homolog A622-like

hetero molecules

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

2

B: Isoflavone reductase homolog A622-like

hetero molecules

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

3

C: Isoflavone reductase homolog A622-like

hetero molecules

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

Unit cell

Length a, b, c (Å)	66.227, 64.917, 115.352
Angle α, β, γ (deg.)	90, 101.513, 90
Int Tables number	4
Space group name H-M	P12₁1

Noncrystallographic symmetry (NCS)

NCS domain:

ID	Ens-ID	Details (eV)
1	1	A
2	1	A
3	2	A
4	2	A
5	3	A
6	3	A

NCS domain segments:

Beg auth comp-ID: GLU / Beg label comp-ID: GLU / End auth comp-ID: VAL / End label comp-ID: VAL / Auth asym-ID: A / Label asym-ID: A / Auth seq-ID: 5 - 310 / Label seq-ID: 5 - 310

Dom-ID	Component-ID	Ens-ID
1	1	1
2	1	1
3	2	2
4	2	2
5	3	3
6	3	3

NCS ensembles :

ID	Details (eV)
1	Local NCS retraints between domains: 1 2
2	Local NCS retraints between domains: 3 4
3	Local NCS retraints between domains: 5 6

#1: Protein	Isoflavone reductase homolog A622-like / NtA622L Mass: 34640.734 Da / Num. of mol.: 3 / Mutation: K99N M201I M259L Source method: isolated from a genetically manipulated source Source: (gene. exp.) Nicotiana tabacum (common tobacco) / Gene: A622L, IRL2 / Production host: Escherichia coli (E. coli) References: UniProt: B6VRE6, Oxidoreductases; Acting on the CH-CH group of donors; With NAD+ or NADP+ as acceptor
#2: Chemical	ChemComp-NDP / NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE Mass: 745.421 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C₂₁H₃₀N₇O₁₇P₃
#3: Chemical	ChemComp-A1JBP / Trigonelline / 1-methylpyridin-1-ium-3-carboxylic acid Mass: 138.144 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C₇H₈NO₂ / Feature type: SUBJECT OF INVESTIGATION
#4: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 105 / Source method: isolated from a natural source / Formula: H₂O
Has ligand of interest	Y
Has protein modification	N

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Experiment

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

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Sample preparation

Crystal	Density Matthews: 2.34 Å³/Da / Density % sol: 47.39 %
Crystal grow	Temperature: 291 K / Method: vapor diffusion, sitting drop / pH: 6.5 Details: 5 mg/ml Protein, 1 mM Trigonelline, 0.5 mM NADPH, 0.1 M Sodium chloride, 0.05 M BIS-TRIS, 12.5% PEG 3350, 500x diluted seed stock

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

ESRF

/ Beamline: ID23-2 / Wavelength: 0.87313 Å

Detector

Type: DECTRIS EIGER X 9M / Detector: PIXEL / Date: Apr 18, 2024

Radiation

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

Radiation wavelength

Wavelength: 0.87313 Å / Relative weight: 1

Reflection

Resolution: 2.6→64.92 Å / Num. obs: 29126 / % possible obs: 97.8 % / Redundancy: 3.3 % / CC_1/2: 0.615 / R_merge(I) obs: 0.267 / R_pim(I) all: 0.266 / R_rim(I) all: 0.377 / Net I/σ(I): 5.9

Reflection shell

Diffraction-ID: 1

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Num. unique obs	CC_1/2	R_pim(I) all	R_rim(I) all
9.01-64.92	2.9	0.098	697	0.777	0.098	0.138
2.6-2.72	3.5	1.026	3585	0.436	1.023	1.449

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0425	refinement
Aimless		data scaling
XDS		data reduction
PHASER		phasing
Coot		model building

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 2.6→56.293 Å / Cor.coef. F_o:F_c: 0.944 / Cor.coef. F_o:F_c free: 0.894 / SU B: 40.134 / SU ML: 0.38 / Cross valid method: FREE R-VALUE / ESU R Free: 0.373
Details: Hydrogens have been added in their riding positions

	R_factor	Num. reflection	% reflection
R_free	0.2678	1471	5.053 %
R_work	0.2062	27639	-
all	0.209	-	-
obs	-	29110	97.609 %

Solvent computation

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

Displacement parameters

B_iso mean: 60.844 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	-2.908 Å²	-0 Å²	1.836 Å²
2-	-	1.061 Å²	-0 Å²
3-	-	-	1.015 Å²

Refinement step

Cycle: LAST / Resolution: 2.6→56.293 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	7251	0	174	105	7530

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.015	0.012	7596
X-RAY DIFFRACTION	r_bond_other_d	0.003	0.016	7272
X-RAY DIFFRACTION	r_angle_refined_deg	2.671	1.828	10338
X-RAY DIFFRACTION	r_angle_other_deg	0.954	1.752	16812
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	7.772	5	915
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	16.29	6.071	42
X-RAY DIFFRACTION	r_dihedral_angle_other_2_deg	7.59	5	6
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	18.261	10	1302
X-RAY DIFFRACTION	r_dihedral_angle_6_deg	15.411	10	318
X-RAY DIFFRACTION	r_chiral_restr	0.13	0.2	1188
X-RAY DIFFRACTION	r_gen_planes_refined	0.012	0.02	8583
X-RAY DIFFRACTION	r_gen_planes_other	0.003	0.02	1620
X-RAY DIFFRACTION	r_nbd_refined	0.21	0.2	1509
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.17	0.2	6775
X-RAY DIFFRACTION	r_nbtor_refined	0.178	0.2	3644
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.082	0.2	4219
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.175	0.2	149
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.272	0.2	31
X-RAY DIFFRACTION	r_nbd_other	0.174	0.2	81
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.322	0.2	5
X-RAY DIFFRACTION	r_mcbond_it	1.822	1.213	3669
X-RAY DIFFRACTION	r_mcbond_other	1.822	1.213	3669
X-RAY DIFFRACTION	r_mcangle_it	2.976	2.183	4581
X-RAY DIFFRACTION	r_mcangle_other	2.976	2.183	4582
X-RAY DIFFRACTION	r_scbond_it	2.044	1.31	3927
X-RAY DIFFRACTION	r_scbond_other	2.043	1.31	3928
X-RAY DIFFRACTION	r_scangle_it	3.316	2.35	5757
X-RAY DIFFRACTION	r_scangle_other	3.316	2.35	5757
X-RAY DIFFRACTION	r_lrange_it	7.512	12.766	8197
X-RAY DIFFRACTION	r_lrange_other	7.512	12.76	8196
X-RAY DIFFRACTION	r_ncsr_local_group_1	0.126	0.05	9490
X-RAY DIFFRACTION	r_ncsr_local_group_2	0.117	0.05	9529
X-RAY DIFFRACTION	r_ncsr_local_group_3	0.123	0.05	9406

Refine LS restraints NCS

Ens-ID	Dom-ID	Auth asym-ID	Refine-ID	Type	Rms dev position (Å)	Weight position
1	1	A	X-RAY DIFFRACTION	Local ncs	0.12552	0.05007
1	2	A	X-RAY DIFFRACTION	Local ncs	0.12552	0.05007
2	3	A	X-RAY DIFFRACTION	Local ncs	0.11677	0.05007
2	4	A	X-RAY DIFFRACTION	Local ncs	0.11677	0.05007
3	5	A	X-RAY DIFFRACTION	Local ncs	0.12272	0.05007
3	6	A	X-RAY DIFFRACTION	Local ncs	0.12272	0.05007

LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Total num. of bins used: 20

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	R_factor all	Num. reflection all	Fsc free	Fsc work	% reflection obs (%)	WR_factor R_work
2.6-2.667	0.348	107	0.306	2063	0.308	2184	0.91	0.947	99.359	0.302
2.667-2.74	0.328	120	0.305	1963	0.306	2114	0.953	0.949	98.5336	0.297
2.74-2.82	0.349	92	0.291	1992	0.294	2104	0.929	0.954	99.0494	0.28
2.82-2.906	0.345	106	0.263	1861	0.267	1987	0.93	0.962	98.9935	0.246
2.906-3.001	0.371	86	0.257	1843	0.262	1957	0.93	0.966	98.5692	0.237
3.001-3.106	0.354	87	0.244	1781	0.249	1893	0.921	0.968	98.6793	0.22
3.106-3.223	0.334	109	0.241	1691	0.246	1834	0.928	0.968	98.1461	0.218
3.223-3.354	0.326	78	0.229	1638	0.233	1743	0.936	0.972	98.4509	0.21
3.354-3.503	0.27	72	0.227	1576	0.229	1678	0.941	0.974	98.2122	0.209
3.503-3.673	0.285	79	0.214	1539	0.217	1637	0.958	0.976	98.8393	0.196
3.673-3.871	0.253	80	0.202	1417	0.205	1530	0.97	0.979	97.8431	0.19
3.871-4.104	0.244	80	0.192	1321	0.195	1446	0.966	0.979	96.888	0.174
4.104-4.386	0.234	88	0.17	1254	0.174	1389	0.967	0.982	96.6163	0.15
4.386-4.735	0.208	64	0.147	1153	0.15	1279	0.973	0.986	95.1525	0.133
4.735-5.183	0.18	43	0.17	1053	0.17	1164	0.979	0.984	94.1581	0.156
5.183-5.789	0.259	52	0.176	986	0.18	1087	0.963	0.984	95.4922	0.163
5.789-6.673	0.216	53	0.173	881	0.175	959	0.972	0.982	97.3931	0.16
6.673-8.144	0.232	36	0.154	732	0.157	812	0.962	0.984	94.5813	0.15
8.144-11.399	0.198	27	0.141	573	0.143	642	0.957	0.985	93.4579	0.141
11.399-56.293	0.25	12	0.24	322	0.241	380	0.965	0.956	87.8947	0.253

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	2.2679	0.2297	-0.3503	3.4873	-0.1252	2.255	0.0934	0.1692	-0.102	-0.1341	-0.0372	0.0072	-0.0269	-0.2447	-0.0562	0.4948	0.0128	0.0013	0.4622	-0.0173	0.0077	6.4601	-1.2139	17.4789
2	1.8942	-0.2317	0.0539	3.7954	0.1729	1.8322	0.0059	0.2519	-0.3087	-0.0467	0.0774	-0.0062	0.0075	-0.125	-0.0833	0.5082	0.0194	0.0061	0.5506	0.0267	0.1095	26.3247	-2.3867	-19.5376
3	2.3423	0.6237	-0.2065	3.8103	-0.2418	2.4527	-0.038	-0.8917	-0.4484	0.165	-0.0717	-0.3877	0.0449	-0.2955	0.1097	0.5634	-0.028	-0.0103	0.9132	0.2316	0.2108	4.4618	-19.089	54.8343

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	ALL	A	5 - 402
2	X-RAY DIFFRACTION	2	ALL	B	5 - 402
3	X-RAY DIFFRACTION	3	ALL	C	5 - 402

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