Solving structures by NMR Sample Preparation Structural restraints .Cloning,expression,purification .NOE,H-bonds .Isotope labelling [5N,13C/15N,[2H/3C/15N ·J-couplings .Residual dipolar couplings,T1/T2 .Chemical shifts Resonance Assignments ·Backbone ·Side chains Structure Calculation .Distance geometry .Restrained molecular dynamics .Simulated annealing Secondary Structure Chemical shift Ensemble of3D structures 3 Solving structures by NMR Sample Preparation •Cloning, expression, purification •Isotope labelling [ 15N], [13C/15N],[ 2H/13C/15N] Resonance Assignments • Backbone • Side chains Secondary Structure Chemical shift Structural restraints •NOE, H-bonds •J-couplings •Residual dipolar couplings, T1/T2 •Chemical shifts Structure Calculation •Distance geometry •Restrained molecular dynamics •Simulated annealing Ensemble of 3D structures 3