2KO1

Solution NMR structure of the ACT domain from GTP pyrophosphokinase of Chlorobium tepidum. Northeast Structural Genomics Consortium Target CtR148A

SOLUTION NMR

NMR Experiment
Experiment	Type	Sample Contents	Solvent	Ionic Strength	pH	Pressure	Temperature (K)	Spectrometer
1	1D 15N T1	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
2	1D 15N T2	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
3	2D 1H-15N HSQC	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
4	2D 1H-13C HSQC aliphatic	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
5	2D 1H-13C CT-HSQC aliphatic	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
6	2D 1H-13C CT-HSQC aromatic	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
7	3D CBCA(CO)NH	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
8	3D HNCACB	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
9	3D HNCO	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
10	3D HN(CA)CO	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
11	3D HBHA(CO)NH	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
12	3D HCCH-COSY	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
13	3D HCCH-TOCSY	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
14	3D 1H-13C NOESY	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
15	3D 1H-15N/13C NOESY	0.3 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
16	2D 1H-13C CT-HSQC methyl	0.5 mM [U-5% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
17	2D 1H-15N HSQC J-modulated	0.5 mM [U-5% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide	90% H2O/10% D2O	225	6.5	ambient	298
18	2D 1H-15N HSQC J-modulated	0.3 mM [U-5% 13C; U-100% 15N] CtR148A, 18 mM MES, 180 mM sodium chloride, 4.5 mM calcium chloride, 9 mM DTT, 45 uM DSS, 0.045 % sodium azide, 4 % polyethylene glycol	84% H2O/16% D2O	225	6.5	ambient	298
19	2D 1H-15N HSQC J-modulated	0.45 mM [U-5% 13C; U-100% 15N] CtR148A, 18 mM MES, 180 mM sodium chloride, 4.5 mM calcium chloride, 9 mM DTT, 45 uM DSS, 0.045 % sodium azide, 7 % polyacrylamide	86% H2O/14% D2O	225	6.5	ambient	298
20	3D 1H-13C X-filtered NOESY	0.5 mM [U-100% 13C; U-100% 15N] CtR148A, 20 mM MES, 200 mM sodium chloride, 5 mM calcium chloride, 10 mM DTT, 50 uM DSS, 0.05 % sodium azide, 0.5 mM CtR148A	90% H2O/10% D2O	225	6.5	ambient	298

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

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, and RDCs from two alignment media. 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	processing	PROSA	6.4	Guntert
3	chemical shift assignment	CARA	1.8.4	Keller and Wuthrich
4	data analysis	CARA	1.8.4	Keller and Wuthrich
5	peak picking	CARA	1.8.4	Keller and Wuthrich
6	chemical shift assignment	PINE	1.0	Bahrami, Markley, Assadi, and Eghbalnia
7	data analysis	TALOS+	1.2009.0721.18	Yang,Cornilescu, Delaglio and Bax
8	structure solution	CYANA	3.0	Guntert, Mumenthaler and Wuthrich
9	refinement	CNS	1.2.1	Brunger, Adams, Clore, Gros, Nilges and Read
10	structure validation	AutoStructure	2.2.1	Huang, Tejero, Powers and Montelione
11	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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2KO1

Solution NMR structure of the ACT domain from GTP pyrophosphokinase of Chlorobium tepidum. Northeast Structural Genomics Consortium Target CtR148A