PDB-9fwd: Structure of indole-3-acetic acid-amido synthetase GH3.6 from A.t...

ID or keywords:

Entry

Database: PDB / ID: 9fwd

Title

Structure of indole-3-acetic acid-amido synthetase GH3.6 from A.thaliana in complex with AMP

Components

Indole-3-acetic acid-amido synthetase GH3.6

Keywords

LIGASE / Plant enzyme / complex / AMP / plant hormone (auxin) inactivation

Function / homology

Function and homology information

indole-3-acetic acid amido synthetase activity / unidimensional cell growth / Ligases; Forming carbon-nitrogen bonds; Acid-amino-acid ligases (peptide synthases) / response to auxin / auxin-activated signaling pathway / cytoplasm
Similarity search - Function

Biological species

Arabidopsis thaliana (thale cress)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.925 Å

Authors

Kopecny, D. / Briozzo, P.

Funding support

Czech Republic,

France, 2items

Citation

Journal: J.Exp.Bot. / Year: 2025
Title: Phenylacetic acid metabolism in land plants: novel pathways and metabolites.
Authors: Hladik, P. / Brunoni, F. / Zukauskaite, A. / Zatloukal, M. / Belicek, J. / Kopecny, D. / Briozzo, P. / Ferchaud, N. / Novak, O. / Pencik, A.

History

Deposition	Jun 29, 2024	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Apr 9, 2025	Provider: repository / Type: Initial release

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	9fwd.cif.gz	682.2 KB	Display	PDBx/mmCIF format
PDB format	pdb9fwd.ent.gz	572 KB	Display	PDB format
PDBx/mmJSON format	9fwd.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

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

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Related structure data

Related structure data	9fxdC C: citing same article (ref.)
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	#129 - Sep 2010 Isocitrate Dehydrogenase similarity (1) #154 - Oct 2012 Citric Acid Cycle similarity (1) #24 - Dec 2001 Glycogen Phosphorylase similarity (1) #55 - Jul 2004 DNA Ligase similarity (1) #16 - Apr 2001 Aminoacyl-tRNA Synthetases similarity (1)

Deposited unit

A: Indole-3-acetic acid-amido synthetase GH3.6

B: Indole-3-acetic acid-amido synthetase GH3.6

hetero molecules

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

1

A: Indole-3-acetic acid-amido synthetase GH3.6

hetero molecules

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

2

B: Indole-3-acetic acid-amido synthetase GH3.6

hetero molecules

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

Unit cell

Length a, b, c (Å)	196.975, 196.975, 65.206
Angle α, β, γ (deg.)	90, 90, 120
Int Tables number	172
Space group name H-M	P6₄

#1: Protein	Indole-3-acetic acid-amido synthetase GH3.6 / Auxin-responsive GH3-like protein 6 / AtGH3-6 / Protein DWARF IN LIGHT 1 / DFL-1 Mass: 70001.453 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Details: N-terminal His-tag / Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Gene: GH3.6, DFL1, At5g54510, F24B18.13 / Plasmid: pETM11 / Production host: Escherichia coli (E. coli) References: UniProt: Q9LSQ4, Ligases; Forming carbon-nitrogen bonds; Acid-amino-acid ligases (peptide synthases)
#2: Chemical	ChemComp-AMP / ADENOSINE MONOPHOSPHATE Mass: 347.221 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₁₀H₁₄N₅O₇P / Feature type: SUBJECT OF INVESTIGATION / Comment: AMP*YM
#3: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 569 / 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: 2.61 Å³/Da / Density % sol: 52.88 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 6.5 Details: 0.1 M MES pH 6.5, 20 % PEG 4000, 0.6 M NaCl; protein at final concentration of 8.2 mg/ml and 10 mM AMP

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

SOLEIL

/ Beamline: PROXIMA 2 / Wavelength: 0.987 Å

Detector

Type: DECTRIS EIGER X 9M / Detector: PIXEL / Date: May 11, 2023

Radiation

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

Radiation wavelength

Wavelength: 0.987 Å / Relative weight: 1

Reflection

Resolution: 1.925→85.293 Å / Num. obs: 58596 / % possible obs: 95.4 % / Redundancy: 20.89 %
Details: Some remarks regarding the mmCIF items written, the PDB Exchange Dictionary (PDBx/mmCIF) Version 5.0 supporting the data files in the current PDB archive (dictionary version 5.325, last ...Details: Some remarks regarding the mmCIF items written, the PDB Exchange Dictionary (PDBx/mmCIF) Version 5.0 supporting the data files in the current PDB archive (dictionary version 5.325, last updated 2020-04-13: http://mmcif.wwpdb.org/dictionaries/mmcif_pdbx_v50.dic/Index/) and the actual quantities provided by MRFANA (https://github.com/githubgphl/MRFANA) from the autoPROC package (https://www.globalphasing.com/autoproc/). In general, the mmCIF categories here should provide items that are currently used in the PDB archive. If there are alternatives, the one recommended by the PDB developers has been selected. The distinction between *_all and *_obs quantities is not always clear: often only one version is actively used within the PDB archive (or is the one recommended by PDB developers). The intention of distinguishing between classes of reflections before and after some kind of observation criterion was applied, can in principle be useful - but such criteria change in various ways throughout the data processing steps (rejection of overloaded or too partial reflections, outlier/misfit rejections during scaling etc) and there is no retrospect computation of data scaling/merging statistics for the reflections used in the final refinement (where another observation criterion might have been applied). Typical data processing will usually only provide one version of statistics at various stages and these are given in the recommended item here, irrespective of the "_all" and "_obs" connotation, see e.g. the use of _reflns.pdbx_Rmerge_I_obs, _reflns.pdbx_Rrim_I_all and _reflns.pdbx_Rpim_I_all. Please note that all statistics related to "merged intensities" (or "merging") are based on inverse-variance weighting of the individual measurements making up a symmetry-unique reflection. This is standard for several decades now, even if some of the dictionary definitions seem to suggest that a simple "mean" or "average" intensity is being used instead. R-values are always given for all symmetry-equivalent reflections following Friedel's law, i.e. Bijvoet pairs are not treated separately (since we want to describe the overall mean intensity and not the mean I(+) and I(-) here). The Rrim metric is identical to the Rmeas R-value and only differs in name. _reflns.pdbx_number_measured_all is the number of measured intensities just before the final merging step (at which point no additional rejection takes place). _reflns.number_obs is the number of symmetry-unique observations, i.e. the result of merging those measurements via inverse-variance weighting. _reflns.pdbx_netI_over_sigmaI is based on the merged intensities (_reflns.number_obs) as expected. _reflns.pdbx_redundancy is synonymous with "multiplicity". The per-shell item _reflns_shell.number_measured_all corresponds to the overall value _reflns.pdbx_number_measured_all. The per-shell item _reflns_shell.number_unique_all corresponds to the overall value _reflns.number_obs. The per-shell item _reflns_shell.percent_possible_all corresponds to the overall value _reflns.percent_possible_obs. The per-shell item _reflns_shell.meanI_over_sigI_obs corresponds to the overall value given as _reflns.pdbx_netI_over_sigmaI. But be aware of the incorrect definition of the former in the current dictionary!
CC_1/2: 0.998 / CC_1/2 anomalous: -0.199 / R_merge(I) obs: 0.1745 / R_pim(I) all: 0.0388 / R_rim(I) all: 0.1788 / AbsDiff over sigma anomalous: 0.774 / B_aniso tensor eigenvalue 1: 60.9 Å² / B_aniso tensor eigenvalue 2: 60.9 Å² / B_aniso tensor eigenvalue 3: 29.1 Å² / B_aniso tensor eigenvector 1 ortho1: 1 / B_aniso tensor eigenvector 1 ortho2: 0 / B_aniso tensor eigenvector 1 ortho3: 0 / B_aniso tensor eigenvector 2 ortho1: 0 / B_aniso tensor eigenvector 2 ortho2: 1 / B_aniso tensor eigenvector 2 ortho3: 0 / B_aniso tensor eigenvector 3 ortho1: 0 / B_aniso tensor eigenvector 3 ortho2: 0 / B_aniso tensor eigenvector 3 ortho3: 1 / Aniso diffraction limit 1: 2.613 Å / Aniso diffraction limit 2: 2.613 Å / Aniso diffraction limit 3: 1.858 Å / Aniso diffraction limit axis 1 ortho1: 1 / Aniso diffraction limit axis 1 ortho2: 0 / Aniso diffraction limit axis 1 ortho3: 0 / Aniso diffraction limit axis 2 ortho1: 0 / Aniso diffraction limit axis 2 ortho2: 1 / Aniso diffraction limit axis 2 ortho3: 0 / Aniso diffraction limit axis 3 ortho1: 0 / Aniso diffraction limit axis 3 ortho2: 0 / Aniso diffraction limit axis 3 ortho3: 1 / Net I/σ(I): 14.56 / Num. measured all: 1224027 / Observed signal threshold: 1.2 / Orthogonalization convention: pdb / % possible anomalous: 95.3 / % possible ellipsoidal: 95.4 / % possible ellipsoidal anomalous: 95.3 / % possible spherical: 53.5 / % possible spherical anomalous: 53.5 / Redundancy anomalous: 10.67 / Signal type: local

Reflection shell

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Mean I/σ(I) obs	Num. measured all	Num. measured obs	Num. unique all	Num. unique obs	CC_1/2	CC_1/2 anomalous	R_pim(I) all	R_rim(I) all	AbsDiff over sigma anomalous	% possible anomalous	% possible ellipsoidal	% possible ellipsoidal anomalous	% possible spherical	% possible spherical anomalous	Redundancy anomalous	% possible all
6.545-85.293	20.14	0.0432	51.4	58960	58960	2927	2927	0.999	-0.528	0.0098	0.0443	0.63	99.7	100	99.7	100	99.7	10.85	100
5.17-6.545	20.02	0.0735	37.15	58727	58727	2933	2933	0.998	-0.34	0.0167	0.0754	0.739	99.6	100	99.6	100	99.6	10.52	100
4.508-5.17	20.24	0.0735	37.06	59313	59313	2931	2931	0.998	-0.282	0.0167	0.0754	0.752	100	100	100	100	100	10.44	100
4.088-4.508	21.35	0.0888	33.35	62440	62440	2925	2925	0.998	-0.26	0.0197	0.091	0.784	100	100	100	100	100	11	100
3.791-4.088	19.08	0.1113	26.13	55975	55975	2933	2933	0.997	-0.172	0.026	0.1144	0.794	99.4	100	99.4	100	99.4	9.95	100
3.565-3.791	19.8	0.1527	20.62	57997	57997	2929	2929	0.996	-0.129	0.0352	0.1568	0.79	99.9	99.9	99.9	99.9	99.9	10.14	99.9
3.384-3.565	20.88	0.2083	16.46	61189	61189	2930	2930	0.994	-0.076	0.0465	0.2135	0.804	99.9	99.9	99.9	99.9	99.9	10.68	99.9
3.233-3.384	21.36	0.2771	13.25	62599	62599	2930	2930	0.99	-0.064	0.0612	0.2838	0.803	99.7	99.8	99.7	99.8	99.7	10.93	99.8
3.108-3.233	21.48	0.3883	9.88	63001	63001	2933	2933	0.983	-0.016	0.0854	0.3977	0.825	100	100	100	100	100	10.98	100
2.999-3.108	21.59	0.4907	8.14	63247	63247	2930	2930	0.977	-0.038	0.1074	0.5024	0.798	99.8	99.8	99.8	99.8	99.8	11.02	99.8
2.906-2.999	21.79	0.6212	6.61	63745	63745	2925	2925	0.97	-0.027	0.1353	0.6359	0.809	99.8	99.8	99.8	99.8	99.8	11.13	99.8
2.82-2.906	22.05	0.7999	5.23	64570	64570	2929	2929	0.943	0.016	0.1734	0.8186	0.794	99.6	99.6	99.6	99.6	99.6	11.24	99.6
2.745-2.82	21.93	0.9056	4.71	64270	64270	2931	2931	0.937	-0.005	0.1966	0.9268	0.783	99.9	99.9	99.9	99.9	99.9	11.19	99.9
2.677-2.745	22.02	1.2113	3.56	64410	64410	2925	2925	0.896	0.031	0.2622	1.2396	0.776	99.4	99	99.4	99	99.4	11.21	99
2.608-2.677	21.93	1.32	3.2	64386	64386	2936	2936	0.865	-0.044	0.2871	1.351	0.754	91.5	89.3	91.5	88.9	91.3	11.05	89.3
2.525-2.608	21.65	1.2363	3.45	63470	63470	2931	2931	0.887	0.007	0.2696	1.2656	0.773	88.9	88.9	88.9	65.6	68.1	10.84	88.9
2.431-2.525	21.68	1.2752	3.31	63497	63497	2929	2929	0.853	0.004	0.2773	1.3051	0.771	91.6	91.6	91.6	49.9	51.8	10.84	91.6
2.322-2.431	21.42	1.4187	2.96	62713	62713	2928	2928	0.844	-0.021	0.3098	1.4523	0.755	91	91	91	36.6	37.9	10.71	91
2.178-2.322	20.05	1.4371	2.82	58741	58741	2929	2929	0.809	-0.006	0.3223	1.4732	0.764	82.9	83	82.9	22.3	23.1	10.03	83
1.925-2.178	17.32	1.7269	2	50777	50777	2932	2932	0.637	-0.032	0.4107	1.7764	0.76	76.6	76.6	76.6	8.7	9	8.66	76.6

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Processing

Software

Name	Version	Classification
MxCuBE		data collection
autoPROC	1.0.5 20211020	data processing
Aimless	0.7.7	data scaling
BUSTER	2.10.4	refinement
XDS	Jun 30, 2023	data reduction
PHASER		phasing
Coot		model building

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 1.925→28.04 Å / Cor.coef. F_o:F_c: 0.928 / Cor.coef. F_o:F_c free: 0.899 / Cross valid method: THROUGHOUT / SU R Blow DPI: 0.355 / SU R_free Blow DPI: 0.233

	R_factor	Num. reflection	% reflection	Selection details
R_free	0.2402	2886	-	RANDOM
R_work	0.2027	-	-	-
obs	0.2046	58553	53.5 %	-

Displacement parameters

B_iso mean: 44.52 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	3.2102 Å²	0 Å²	0 Å²
2-	-	3.2102 Å²	0 Å²
3-	-	-	-6.4204 Å²

Refine analyze

Luzzati coordinate error obs: 0.28 Å

Refinement step

Cycle: LAST / Resolution: 1.925→28.04 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	9301	0	46	569	9916

Refine LS restraints

Refine-ID	Type	Dev ideal	Number	Restraint function	Weight
X-RAY DIFFRACTION	t_bond_d	0.008	18812	HARMONIC	2
X-RAY DIFFRACTION	t_angle_deg	1.01	34028	HARMONIC	2
X-RAY DIFFRACTION	t_dihedral_angle_d		5619	SINUSOIDAL	2
X-RAY DIFFRACTION	t_gen_planes		2911	HARMONIC	5
X-RAY DIFFRACTION	t_it		18812	HARMONIC	10
X-RAY DIFFRACTION	t_chiral_improper_torsion		1249	SEMIHARMONIC	5
X-RAY DIFFRACTION	t_ideal_dist_contact		16421	SEMIHARMONIC	4
X-RAY DIFFRACTION	t_omega_torsion	3.28
X-RAY DIFFRACTION	t_other_torsion	16.28

LS refinement shell

Resolution: 1.93→2.06 Å

	R_factor	Num. reflection	% reflection
R_free	0.2879	63	-
R_work	0.2573	-	-
obs	0.2589	1172	5.78 %

Refinement TLS params.

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	0.7818	-0.1017	-0.458	0.5447	0.1214	2.3999	0.0233	-0.0103	-0.0972	-0.0103	-0.0201	0.0499	-0.0972	0.0499	-0.0031	-0.0578	0.073	-0.0345	-0.0287	-0.0316	-0.1673	-38.9373	-42.425	-1.8623
2	0.4799	-0.0528	0.1979	0.5139	-0.1971	1.7184	-0.0103	-0.0379	-0.2112	-0.0379	0.0123	-0.0938	-0.2112	-0.0938	-0.002	0.0684	0.0547	0.004	-0.0712	0.0112	-0.1644	-18.5339	-87.8025	18.3382

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	{ A\|* }	A	13 - 609
2	X-RAY DIFFRACTION	1	{ A\|* }	A	701
3	X-RAY DIFFRACTION	2	{ B\|* }	B	13 - 610
4	X-RAY DIFFRACTION	2	{ B\|* }	B	701

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