Solution Structure Determination by NMR and Distance Geometry Calculations 1. Pure Protein (0.3 mL, mM; ~ 10 mg) 2. Sequence-specific resonance assignments 3. Sequence-specifice distance constraints disulfide bonds spin-spin coupling constants internuclear distance constraints from NOEs residual dipolar couplings hydrogen bonds standard bond lengths; bond angles 4. Distance Geometry Calculations 5. Constrained Energy Minimization; Refinement

Basic Principles Nuclear Spin and Magnetization

Magnetization and Coherence

Fourier Transform NMR Ernst & Anderson Rev. Scientific Instrumentation 37: 93 (1966)

Nuclear Relaxation

Spin-Spin Coupling

Chemical Shift with Spin-Spin Coupling

1D NMR Spectrum of a Protein

2D Correlated SpectroscopY

AV-COSY

Nuclear Overhauser Effect (NOE) Def: The change in the intensity of resonance i upon irradiation of resonance j due to “cross relaxation” between spins i and j through space

2D NOESY Spectrum of a Protein

Some Useful 2D NMR Experiments

1 H- 13 C Heteronuclear Correlation Spectrum

Solution Structure Determination by NMR and Distance Geometry Calculations 1. Pure Protein (0.3 mL, mM; ~ 10 mg) 2. Sequence-specific resonance assignments 3. Sequence-specifice distance constraints disulfide bonds spin-spin coupling constants internuclear distance constraints from NOEs residual dipolar couplings hydrogen bonds standard bond lengths; bond angles 4. Distance Geometry Calculations 5. Constrained Energy Minimization; Refinement

WR41 Protein NMR Structures C-TmZip ER14 MMP-1 IL13 FGF-2 WR90 WR64 LC8 ER115 N-TmZip JR19 ZR18 OP3 WR33 ZR31 ER75 Z-domain IR24

ER75: Ecol YgdK (nmr) ER30: Ecol SufE (xray) IR24: Hinf IscU (nmr) SR17 Bsub IscU (nmr) N N CC YgdK G. Liu et al. (Szyperski) Gram neg IscU IscU, SufE,YgdK Domain Families are Structurally Similar YgdK cf SufE A. Kuzin et al. (Hunt) Ramelot et al, J. Mol. Biol Goldsmith et al., J. Mol. Biol Liu et al. submitted T. Ramelot et al. (Kennedy) Gram pos IscU G. Kornhaber et al. (Montelione)