2JUN

Structure of the MID1 tandem B-boxes reveals an interaction reminiscent of intermolecular RING heterodimers

SOLUTION NMR

NMR Experiment
Experiment	Type	Sample Contents	Solvent	Ionic Strength	pH	Pressure	Temperature (K)	Spectrometer
1	2D 1H-15N HSQC	0.75-1.0 mM 15N & 15N/13C labeled protein in 50 mM Tris-HCl, 300 mM NaCl, 10 mM 2-mercaptoethanol, 5 mM ZnCl2, 2 % azide, and 0.015% Triton-X 100, pH 7.5, 90% H2O, 10% D2O	90% H2O/10% D2O		7.5	ambient	294
2	3D HNCA	0.75-1.0 mM 15N & 15N/13C labeled protein in 50 mM Tris-HCl, 300 mM NaCl, 10 mM 2-mercaptoethanol, 5 mM ZnCl2, 2 % azide, and 0.015% Triton-X 100, pH 7.5, 90% H2O, 10% D2O	90% H2O/10% D2O		7.5	ambient	294
3	3D HNCACB	0.75-1.0 mM 15N & 15N/13C labeled protein in 50 mM Tris-HCl, 300 mM NaCl, 10 mM 2-mercaptoethanol, 5 mM ZnCl2, 2 % azide, and 0.015% Triton-X 100, pH 7.5, 90% H2O, 10% D2O	90% H2O/10% D2O		7.5	ambient	294
4	3D 1H-15N NOESY	0.75-1.0 mM 15N & 15N/13C labeled protein in 50 mM Tris-HCl, 300 mM NaCl, 10 mM 2-mercaptoethanol, 5 mM ZnCl2, 2 % azide, and 0.015% Triton-X 100, pH 7.5, 90% H2O, 10% D2O	90% H2O/10% D2O		7.5	ambient	294
5	3D 1H-13C NOESY	0.75-1.0 mM 15N & 15N/13C labeled protein in 50 mM Tris-HCl, 300 mM NaCl, 10 mM 2-mercaptoethanol, 5 mM ZnCl2, 2 % azide, and 0.015% Triton-X 100, pH 7.5, 90% H2O, 10% D2O	90% H2O/10% D2O		7.5	ambient	294
6	3D HCCH-TOCSY	0.75-1.0 mM 15N & 15N/13C labeled protein in 50 mM Tris-HCl, 300 mM NaCl, 10 mM 2-mercaptoethanol, 5 mM ZnCl2, 2 % azide, and 0.015% Triton-X 100, pH 7.5, 90% H2O, 10% D2O	90% H2O/10% D2O		7.5	ambient	294

NMR Spectrometer Information
Spectrometer	Manufacturer	Model	Field Strength
1	Varian	INOVA	600

NMR Refinement
Method	Details	Software
torsion angle dynamics		NMRPipe

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

Computation: NMR Software
#	Classification	Version	Software Name	Author
1	chemical shift calculation	NMRPipe		Delaglio, Grzesiek, Vuister, Zhu, Pfeifer and Bax
2	processing	NMRPipe		Delaglio, Grzesiek, Vuister, Zhu, Pfeifer and Bax
3	chemical shift assignment	Sparky	3.110	Goddard
4	peak picking	Sparky	3.110	Goddard
5	data analysis	TALOS		Cornilescu, Delaglio and Bax
6	refinement	CYANA	2.1	Guntert, Mumenthaler and Wuthrich

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.