Transferred NOE Experiments K A A-B tr NOE NOE r NOE A-B NOE relayed via spin diffusion Direct NOE (A.-B) Mixing time )ms Ni&Scheraga Acc.Chem.Res.27257-264 Transferred NOEs 29cme6tgandmawacaedsocaeaevnemg -NOE build-up curves are used to optimize the mixing time Intermolecular NOEs may also be observed on rates(aso time) -53 Transferred NOE Experiments Ni & Scheraga Acc. Chem. Res. 27 257-264 Ab Bb Af Bf Koff` kon Z Mixing time NOE 600 ms A-B tr NOE Intermolecular tr NOE (Z-B) A-B NOE relayed via spin diffusion Direct NOE (Ab-Bb) 100% 0% 4 types of NOE are possible Transferred NOEs •Ligand must be in large excess over protein - at least 10X, so the protein is saturated with ligand but the linewidths are dominated by free ligand •The off-rate must be fast: the ligand must associate/dissociate a few times during the NOE mixing time •NOE build-up curves are used to optimize the mixing time •Intermolecular NOEs may also be observed •Transferred cross correlated relaxation rates (also depend on correlation time) (Blommers et al 1999 JACS 121 1949-53 Carlomagno 1999 JACS 121 1945-48) •Transferred dipolar couplings have also been observed (special case if protein is weakly aligned) (Koenig et al 2000 J.Biomol. NMR 16 121-125)