2KI8

Solution NMR structure of tungsten formylmethanofuran dehydrogenase subunit D from Archaeoglobus fulgidus, Northeast Structural Genomics Consortium target AtT7

SOLUTION NMR

NMR Experiment
Experiment	Type	Sample Contents	Solvent	Ionic Strength	pH	Pressure	Temperature (K)	Spectrometer
1	2D 1H-15N HSQC	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
2	2D 1H-13C HSQC aliphatic	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
3	2D 1H-13C CT-HSQC aliphatic	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
4	2D 1H-13C CT-HSQC aromatic	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
5	3D HNCO	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
6	3D HN(CA)CO	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
7	3D CBCA(CO)NH	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
8	3D HNCACB	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
9	3D HBHA(CO)NH	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
10	(4,3)D GFT HCCH-COSY aliphatic	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
11	3D HCCH-TOCSY aliphatic	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
12	(4,3)D GFT HCCH-COSY aromatic	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
13	3D 1H-15N,13C NOESY	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
14	3D 1H-13C NOESY	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
15	2D MEXICO	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
16	2D CLEANEX	0.6 mM [U-100% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
17	2D 1H-13C CT-HSQC methyl	0.6 mM [U-7% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
18	2D 1H-15N HSQC	0.6 mM [U-7% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
19	2D 1H-15N TROSY	0.6 mM [U-7% 13C; U-100% 15N] AtT7 protein, 10 mM TRIS, 300 mM sodium chloride, 1 mM benzamidine	90% H2O/10% D2O	300	7.0	ambient	298
20	2D 1H-15N HSQC	0.4 mM [U-7% 13C; U-100% 15N] AtT7 protein, 6.6 mM TRIS, 200 mM sodium chloride, 0.7 mM benzamidine, 13.25 g/L Pf1 phage	86% H2O/14% D2O	200	7.0	ambient	298
21	2D 1H-15N TROSY	0.4 mM [U-7% 13C; U-100% 15N] AtT7 protein, 6.6 mM TRIS, 200 mM sodium chloride, 0.7 mM benzamidine, 13.25 g/L Pf1 phage	86% H2O/14% D2O	200	7.0	ambient	298

NMR Spectrometer Information
Spectrometer	Manufacturer	Model	Field Strength
1	Varian	INOVA	750
2	Varian	INOVA	600
3	Bruker	AVANCE	800

NMR Refinement
Method	Details	Software
simulated annealing	Structure determination was performed iteratively with CYANA v2.1 and CYANA v3.0 using NOE-based constraints, PHI and PSI dihedral angle constraints from TALOS, hydrogen bond constraints based on preliminary structures and CLEANEX/MEXICO data, and RDCs from a single alignment medium. The 20 conformers out of 100 with the lowest target function were further refined by simulated annealing in explicit water bath using the program CNS with PARAM19 force field.	VnmrJ

NMR Ensemble Information
Conformer Selection Criteria	target function
Conformers Calculated Total Number	100
Conformers Submitted Total Number	20
Representative Model	1 (lowest energy)

Computation: NMR Software
#	Classification	Version	Software Name	Author
1	collection	VnmrJ	2.1B	Varian
2	collection	TopSpin		Bruker Biospin
3	processing	TopSpin		Bruker Biospin
4	processing	PROSA	6.0.2	Guntert
5	data analysis	CARA	1.8.4	Keller and Wuthrich
6	peak picking	CARA	1.8.4	Keller and Wuthrich
7	chemical shift assignment	CARA	1.8.4	Keller and Wuthrich
8	chemical shift assignment	AutoAssign	2.3.0	Zimmerman, Moseley, Kulikowski and Montelione
9	data analysis	CSI	2.0	Wishart and Sykes
10	data analysis	TALOS	2007.068.09.07	Shen, Cornilescu, Delaglio and Bax
11	structure solution	CYANA	3.0	Guntert, Mumenthaler and Wuthrich
12	refinement	CNS	1.2.1	Brunger, Adams, Clore, Gros, Nilges and Read
13	structure validation	AutoStructure	2.1.1	Huang, Tejero, Powers and Montelione
14	structure validation	PSVS	1.3	Bhattacharya and Montelione

RCSB PDB Core Operations are funded by the National Science Foundation (DBI-1832184), the US Department of Energy (DE-SC0019749), and the National Cancer Institute, National Institute of Allergy and Infectious Diseases, and National Institute of General Medical Sciences of the National Institutes of Health under grant R01GM133198.

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2KI8

Solution NMR structure of tungsten formylmethanofuran dehydrogenase subunit D from Archaeoglobus fulgidus, Northeast Structural Genomics Consortium target AtT7