Experiment: 2HAC

SOLUTION NMR

NMR Experiment
Experiment	Type	Sample Contents	Solvent	Ionic Strength	pH	Pressure	Temperature (K)	Spectrometer
1	gradient-enhanced HNCA	0.5-1.0 mM disulfide crosslinked zeta-zeta dimer U-15N,U-13C,85%-2H. Sodium dodecyl sulfate (40 mM) and dodecyl phosphocholine (200 mM), 95% H2O, 5% D2O.	95% H2O/5% D2O	20 mM sodium phosphate buffer	7.0	ambient	303
2	3D_15N-separated_NOESY	0.5-1.0 mM disulfide crosslinked zeta-zeta dimer U-15N,U-13C. Sodium dodecyl sulfate (40 mM) and dodecyl phosphocholine (200 mM), 95% H2O, 5% D2O.	95% H2O/5% D2O	20 mM sodium phosphate buffer	7.0	ambient	303
3	3D_15N-separated_NOESY	0.5-1.0 mM disulfide crosslinked zeta-zeta dimer U-15N,U-13C. Sodium dodecyl sulfate (40 mM) and dodecyl phosphocholine (200 mM), 95% H2O, 5% D2O.	95% H2O/5% D2O	20 mM sodium phosphate buffer	7.0	ambient	303
4	3D_13C-separated_NOESY	0.5-1.0 mM disulfide crosslinked zeta-zeta dimer U-15N,U-13C. Sodium dodecyl sulfate (40 mM) and dodecyl phosphocholine (200 mM), 95% H2O, 5% D2O.	95% H2O/5% D2O	20 mM sodium phosphate buffer	7.0	ambient	303
5	3D_15N-separated_NOESY	0.5-1.0 mM disulfide crosslinked zeta-zeta dimer, mixed-label. One monomer: U-15N,U-2H. Other monomer: natural abundance isotopes. Sodium dodecyl sulfate (40 mM) and dodecyl phosphocholine (200 mM), 95% H2O, 5% D2O.	95% H2O/5% D2O	20 mM sodium phosphate buffer	7.0	ambient	303
6	standard HSQC and trosy-HSQC for D(NH) couplings; 2D (CA)CONH quantitative JCH experiment for D(CaHa) couplings	0.5-1.0 mM disulfide crosslinked zeta-zeta dimer U-15N,U-13C. Sodium dodecyl sulfate (40 mM) and dodecyl phosphocholine (200 mM), 95% H2O, 5% D2O. 5% polyacrylamide gel.	95% H2O/5% D2O	20 mM sodium phosphate buffer	7.0	ambient	303

NMR Spectrometer Information
Spectrometer	Manufacturer	Model	Field Strength
1	Bruker	AVANCE	500
2	Bruker	AVANCE	600

NMR Refinement
Method	Details	Software
simulated annealing	426 intramolecular NOE restraints, 46 intermolecular NOE restraints, 42 dihedral angle restraints, 70 residual dipolar coupling restraints.	NMRPipe

NMR Ensemble Information
Conformer Selection Criteria	structures with the lowest energy
Conformers Calculated Total Number	30
Conformers Submitted Total Number	15
Representative Model	1 (closest to the average)

Computation: NMR Software
#	Classification	Version	Software Name	Author
1	processing	NMRPipe	1.0	Frank Delaglio
2	data analysis	CARA	1.5	Rochus Keller
3	refinement	XPLOR-NIH	2.11	Charles Schwieters

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.