2LCB

Solution Structure of a Minor and Transiently Formed State of a T4 Lysozyme Mutant

SOLUTION NMR

NMR Experiment
Experiment	Type	Sample Contents	Solvent	Ionic Strength	pH	Pressure	Temperature (K)	Spectrometer
1	15N CT Trosy CPMG	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
2	15N CT Trosy CPMG	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
3	1H CT CPMG	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
4	1H CT CPMG	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
5	1H ZQ/DQ CPMG	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
6	1H ZQ/DQ CPMG	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
7	13C' CT CPMG	1.5 mM [U-13C; U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
8	13C' CT CPMG	1.5 mM [U-13C; U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
9	13Ca CT CPMG	1.5 mM [U-13Ca; U-15N] T4 L99A	100% D2O	85	5.5	ambient	298
10	13Ca CT CPMG	1.5 mM [U-13Ca; U-15N] T4 L99A	100% D2O	85	5.5	ambient	298
11	1Ha CT CPMG	1.5 mM [U-13C; U-15N; U-50% 2H] T4 L99A	100% D2O	85	5.5	ambient	298
12	1Ha CT CPMG	1.5 mM [U-13C; U-15N; U-50% 2H] T4 L99A	100% D2O	85	5.5	ambient	298
13	Gly 13Ca CT CPMG	1.5 mM [U-13C; U-15N; U-50% 2H] T4 L99A	100% D2O	85	5.5	ambient	298
14	Gly 1Ha CT CPMG	1.5 mM [U-13C; U-15N; U-50% 2H] T4 L99A	100% D2O	85	5.5	ambient	298
15	Gly 1Ha CT CPMG	1.5 mM [U-13C; U-15N; U-50% 2H] T4 L99A	100% D2O	85	5.5	ambient	298
16	1H-15N ZZ exchange	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
17	1H-13C Met ZZ exchange	1.5 mM [ U-15N] 13CH3 Met T4 L99A	100% D2O	85	5.5	ambient	298
18	2D 1H-15N HSQC	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
19	2D 1H-15N HSQC	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
20	2D 1H-15N HMQC	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
21	2D 1H-15N HMQC	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
22	3D HNCO	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
23	3D HNCO	1.5 mM [U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
24	3D MQ HNCO	1.5 mM [U-13C; U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
25	3D MQ HNCO	1.5 mM [U-13C; U-15N; U-2H] T4 L99A	90% H2O/10% D2O	85	5.5	ambient	298
26	2D 1H-13C HSQC	1.5 mM [U-13Ca; U-15N] T4 L99A	100% D2O	85	5.5	ambient	298
27	2D 1H-13C HSQC	1.5 mM [U-13Ca; U-15N] T4 L99A	100% D2O	85	5.5	ambient	298
28	2D 1H-13C HSQC	1.5 mM [U-13C; U-15N; U-50% 2H] T4 L99A	100% D2O	85	5.5	ambient	298
29	2D 1H-13C HSQC	1.5 mM [U-13C; U-15N; U-50% 2H] T4 L99A	100% D2O	85	5.5	ambient	298

NMR Spectrometer Information
Spectrometer	Manufacturer	Model	Field Strength
1	Varian	INOVA	500
2	Varian	INOVA	600
3	Varian	INOVA	800

NMR Refinement
Method	Details	Software
distance geometry, simulated annealing	CS-Rosetta Loop Building	NMRPipe

NMR Ensemble Information
Conformer Selection Criteria	Two Step Selection Criteria based on chemical shift score and Rosetta energy
Conformers Calculated Total Number	9600
Conformers Submitted Total Number	10
Representative Model	1 (lowest energy)

Computation: NMR Software
#	Classification	Version	Software Name	Author
1	processing	NMRPipe		Delaglio, Grzesiek, Vuister, Zhu, Pfeifer and Bax
2	peak picking	Sparky		Goddard
3	extract peak intensities	FuDA		Flemming Hansen
4	cpmg data analysis	CATIA		Flemming Hansen
5	structure solution	CS-ROSETTA		Yang Shen, Robert Vernon, David Baker and Ad Bax
6	refinement	CS-ROSETTA		Yang Shen, Robert Vernon, David Baker and Ad Bax

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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2LCB

Solution Structure of a Minor and Transiently Formed State of a T4 Lysozyme Mutant