Experiment: 2N1R

SOLUTION NMR

NMR Experiment
Experiment	Type	Sample Contents	Solvent	Ionic Strength	pH	Pressure	Temperature (K)	Spectrometer
1	15N-1HN HSQC	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L [U-99% 15N] ammonium chloride, 4 g/L glucose, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L ammonium chloride, 4 g/L glucose, 0.5 mg/mL [U-100% 13C] MYRISTIC ACID, 90% H2O, 10% D2O	90% H2O/10% D2O	150mM NaCl	7	ambient, 1atm atm	293
2	13C-1HN HNCA	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L [U-99% 15N] ammonium chloride, 4 g/L glucose, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L ammonium chloride, 4 g/L glucose, 0.5 mg/mL [U-100% 13C] MYRISTIC ACID, 90% H2O, 10% D2O	90% H2O/10% D2O	150mM NaCl	7	ambient, 1atm atm	293
3	13C-1HN HN(CO)CA	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L [U-99% 15N] ammonium chloride, 4 g/L glucose, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L ammonium chloride, 4 g/L glucose, 0.5 mg/mL [U-100% 13C] MYRISTIC ACID, 90% H2O, 10% D2O	90% H2O/10% D2O	150mM NaCl	7	ambient, 1atm atm	293
4	CBCANH	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L [U-99% 15N] ammonium chloride, 4 g/L glucose, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L ammonium chloride, 4 g/L glucose, 0.5 mg/mL [U-100% 13C] MYRISTIC ACID, 90% H2O, 10% D2O	90% H2O/10% D2O	150mM NaCl	7	ambient, 1atm atm	293
5	CBCA(CO)NH	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L [U-99% 15N] ammonium chloride, 4 g/L glucose, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L ammonium chloride, 4 g/L glucose, 0.5 mg/mL [U-100% 13C] MYRISTIC ACID, 90% H2O, 10% D2O	90% H2O/10% D2O	150mM NaCl	7	ambient, 1atm atm	293
6	CARBON-NITROGEN NOESY (CNNOE)	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L [U-99% 15N] ammonium chloride, 4 g/L glucose, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L ammonium chloride, 4 g/L glucose, 0.5 mg/mL [U-100% 13C] MYRISTIC ACID, 90% H2O, 10% D2O	90% H2O/10% D2O	150mM NaCl	7	ambient, 1atm atm	293
7	15N EDITED 3D NOESY	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L [U-99% 15N] ammonium chloride, 4 g/L glucose, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 1 g/L ammonium chloride, 4 g/L glucose, 0.5 mg/mL [U-100% 13C] MYRISTIC ACID, 90% H2O, 10% D2O	90% H2O/10% D2O	150mM NaCl	7	ambient, 1atm atm	293
8	1H-13C HMQC	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L [U-100% 13C] glucose, 1 g/L ammonium chloride, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L [U-100% 13C] glucose, 1 g/L [U-99% 15N] ammonium chloride, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L glucose, 1 g/L ammonium chloride, 0.5 mg/mL MYRISTIC ACID	100% D2O	150mM NaCl	7	ambient, 1atm atm	293
9	CARBON CARBON NOSEY (CCNOE)	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L [U-100% 13C] glucose, 1 g/L ammonium chloride, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L [U-100% 13C] glucose, 1 g/L [U-99% 15N] ammonium chloride, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L glucose, 1 g/L ammonium chloride, 0.5 mg/mL MYRISTIC ACID	100% D2O	150mM NaCl	7	ambient, 1atm atm	293
10	1H-1H NOESY	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L [U-100% 13C] glucose, 1 g/L ammonium chloride, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L [U-100% 13C] glucose, 1 g/L [U-99% 15N] ammonium chloride, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L glucose, 1 g/L ammonium chloride, 0.5 mg/mL MYRISTIC ACID	100% D2O	150mM NaCl	7	ambient, 1atm atm	293
11	13C-1H HMQC NOESY	50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L [U-100% 13C] glucose, 1 g/L ammonium chloride, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L [U-100% 13C] glucose, 1 g/L [U-99% 15N] ammonium chloride, 0.5 mg/mL MYRISTIC ACID, 50 mM sodium phosphate, 150 mM sodium chloride, 10 mM DTT, 4 g/L glucose, 1 g/L ammonium chloride, 0.5 mg/mL MYRISTIC ACID	100% D2O	150mM NaCl	7	ambient, 1atm atm	293

NMR Spectrometer Information
Spectrometer	Manufacturer	Model	Field Strength
1	Bruker	DMX	800
2	Bruker	DMX	600

NMR Refinement
Method	Details	Software
CYANA	Structures were calculated in torsion angle space with CYANA (http://www.las.jp/index_eg.html) starting from random initial angles. Upper interproton distance bounds of 2.7, 3.3 and 5.0 aE (with appropriate corrections for pseudoatoms) were employed for NOE cross peaks of strong, medium, and weak intensity respectively, which were qualitatively determined following intensity normalization of the different NOE data sets. Residual dipolar couplings were prohibited due to precipitation of the sample. No backbone hydrogen bond or chemical shift-based torsion angle restraints were employed.	NMRView

NMR Ensemble Information
Conformer Selection Criteria	20 structures for lowest energy
Conformers Calculated Total Number	200
Conformers Submitted Total Number	20
Representative Model	1 (fewest violations)

Computation: NMR Software
#	Classification	Version	Software Name	Author
1	chemical shift assignment	NMRView		Johnson, One Moon Scientific
2	data analysis	NMRView		Johnson, One Moon Scientific
3	peak picking	NMRView		Johnson, One Moon Scientific
4	structure calculation	CYANA		Guntert, Mumenthaler and Wuthrich
5	processing	NMRPipe		Delaglio, Grzesiek, Vuister, Zhu, Pfeifer and Bax
6	processing	NMRDraw		Delaglio, Grzesiek, Vuister, Zhu, Pfeifer and Bax
7	collection	TopSpin		Bruker Biospin
8	refinement	CYANA

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.