2LDK

Solution NMR Structure of Protein AAur_3427 from Arthrobacter aurescens, Northeast Structural Genomics Consortium Target AaR96

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] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
2	2D 1H-13C CT-HSQC aliphatic	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
3	3D HNCO	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
4	3D CBCA(CO)NH	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
5	3D HNCACB	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
6	2D 1H-13C CT-HSQC aromatic	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
7	3D simutaneous 13C-aromatic,13C-aliphatic,15N edited 1H-1H NOESY	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
8	3D (H)CCH-TOCSY aliphatic	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
9	3D HBHA(CO)NH	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
10	3D HN(CA)CO	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
11	2D 1H-15N LR-HSQC histidine	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
12	2D (HB)CB(CGCDCE)HDHE	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
13	(4,3)D GFT HCCH-COSY aliphatic	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
14	3D HCCH-COSY aromatic	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
15	2D 1H-13C CT-HSQC methyl	1.7 mM [5% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
16	1D 15N T1	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
17	1D 15N T2	0.6 mM [U-100% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
18	2D J-mod 1H-15N HSQC	1.7 mM [5% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide	90% H2O/10% D2O		6.5	ambient	298
19	2D J-mod 1H-15N HSQC	1.2 mM [5% 13C; U-100% 15N] AaR96, 20 mM MES, 100 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.02 % sodium azide, 4 % C12E5 PEG, 1.3 % hexanol	80% H2O/20% D2O		6.5	ambient	298
20	2D J-mod 1H-15N HSQC	1.1 mM [5% 13C; U-100% 15N] AaR96, 13 mM MES, 70 mM sodium chloride, 3 mM calcium chloride, 10 mM DTT, 33 uM DSS, 0.02 % sodium azide, 12.5 g/l Pf1 phage	80% H2O/20% D2O		6.5	ambient	298

NMR Spectrometer Information
Spectrometer	Manufacturer	Model	Field Strength
1	Bruker	AVANCE	900
2	Varian	INOVA	600
3	Varian	INOVA	750
4	Varian	INOVA	600
5	Bruker	AVANCE	700

NMR Refinement
Method	Details	Software
simulated annealing	Structure determination was performed by running CYANA and AUTOSTRUCTURE in parallel using NOE-based constraints and PHI and PSI dihedral angle constraints from TALOS+. Consensus peak assignments were selected and used in iterative refinement with CYANA, with RDC constraints added at later stages. 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	CNS

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	refinement	CNS	1.2	Brunger, Adams, Clore, Gros, Nilges and Read
2	structure solution	CNS	1.2	Brunger, Adams, Clore, Gros, Nilges and Read
3	geometry optimization	CNS	1.2	Brunger, Adams, Clore, Gros, Nilges and Read
4	refinement	CYANA	3.0	Guntert, Mumenthaler and Wuthrich
5	geometry optimization	CYANA	3.0	Guntert, Mumenthaler and Wuthrich
6	structure solution	CYANA	3.0	Guntert, Mumenthaler and Wuthrich
7	data analysis	AutoStructure	2.2.1	Huang, Tejero, Powers and Montelione
8	refinement	AutoStructure	2.2.1	Huang, Tejero, Powers and Montelione
9	data analysis	AutoAssign	2.3.0	Zimmerman, Moseley, Kulikowski and Montelione
10	chemical shift assignment	AutoAssign	2.3.0	Zimmerman, Moseley, Kulikowski and Montelione
11	processing	NMRPipe		Delaglio, Grzesiek, Vuister, Zhu, Pfeifer and Bax
12	data analysis	XEASY	1.3.13	Bartels et al.
13	collection	TopSpin	2.1	Bruker Biospin
14	processing	TopSpin	2.1	Bruker Biospin
15	collection	VnmrJ	2.2D	Varian
16	geometry optimization	TALOS+	1.2009.0721.18	Shen, Cornilescu, Delaglio and Bax
17	data analysis	CARA	1.8.4	Keller and Wuthrich
18	chemical shift assignment	CARA	1.8.4	Keller and Wuthrich
19	peak picking	CARA	1.8.4	Keller and Wuthrich
20	processing	PROSA	6.4	Guntert
21	refinement	CNS		Brunger, Adams, Clore, Gros, Nilges and Read

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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2LDK

Solution NMR Structure of Protein AAur_3427 from Arthrobacter aurescens, Northeast Structural Genomics Consortium Target AaR96