5O7J

Structural insights into the periplasmic sensor domain of the GacS histidine kinase controlling biofilm formation in Pseudomonas aeruginosa

SOLUTION NMR

NMR Experiment
Experiment	Type	Sample Contents	Solvent	Ionic Strength	pH	Pressure	Temperature (K)	Spectrometer
1	2D 1H-1H TOCSY	0.8 mM [U-99% 15N] GacS periplasmic detection domain, 150 mM sodium chloride, 50 mM sodium phosphate	90% H2O/10% D2O	0.2 M	7	1 atm	298	Bruker AVANCE 600
2	2D 1H-1H NOESY	0.8 mM [U-99% 15N] GacS periplasmic detection domain, 150 mM sodium chloride, 50 mM sodium phosphate	90% H2O/10% D2O	0.2 M	7	1 atm	298	Bruker AVANCE 600
3	2D 1H-15N HSQC	0.8 mM [U-99% 15N] GacS periplasmic detection domain, 150 mM sodium chloride, 50 mM sodium phosphate	90% H2O/10% D2O	0.2 M	7	1 atm	298	Bruker AVANCE 600
4	3D 1H-15N NOESY	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Bruker AVANCE 600
5	3D HNHA	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
6	2D 1H-15N HSQC	0.8 mM [U-99% 15N] GacS periplasmic detection domain, 150 mM sodium chloride, 50 mM sodium phosphate	90% H2O/10% D2O	0.2 M	7	1 atm	298	Bruker AVANCE 600
7	2D 1H-13C HSQC	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Bruker AVANCE 600
8	3D HNCO	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
9	3D HCACO	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
10	3D HNCA	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
14	3D HN(CO)CA	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
13	3D HNCACB	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
12	3D CBCA(CO)NH	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
11	3D HCCH-TOCSY	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
16	3D 1H-13C NOESY	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
15	3D 1H-13C NOESY aromatic	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Varian Uniform NMR System 850
17	2D CBCACO	0.8 mM [U-99% 13C; U-99% 15N] GacS periplasmic detection domain, 50 mM sodium phosphate, 150 mM sodium chloride	90% H2O/10% D2O	0.2 M	7	1 atm	298	Bruker AVANCE 600

NMR Spectrometer Information
Spectrometer	Manufacturer	Model	Field Strength
1	Bruker	AVANCE	600
2	Varian	Uniform NMR System	850

NMR Refinement
Method	Details	Software
simulated annealing		Amber

NMR Ensemble Information
Conformer Selection Criteria	all calculated structures submitted
Conformers Calculated Total Number	20
Conformers Submitted Total Number	20
Representative Model	1 (lowest energy)

Computation: NMR Software
#	Classification	Version	Software Name	Author
1	processing	TopSpin	2.0	Bruker Biospin
2	peak picking	CARA		Keller and Wuthrich
3	chemical shift assignment	CARA		Keller and Wuthrich
4	structure calculation	CYANA		Guntert, Mumenthaler and Wuthrich
5	refinement	Amber		Case, Darden, Cheatham III, Simmerling, Wang, Duke, Luo, ... and Kollman

RCSB PDB Core Operations are funded by the U.S. National Science Foundation (DBI-2321666), 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.