The use of dipolar couplings for determining the solution structure of ratapo-S100B(beta beta)

The relative orientations of adjacent structural elements without many well -defined NOE contacts between them are typically poorly defined in NMR stru ctures. For apo-S100B(PP) and the structurally homologous protein calcyclin , the solution structures determined by conventional NMR exhibited consider able differences and made it impossible to draw unambiguous conclusions reg arding the Ca2+-induced conformational change required for target protein b inding. The structure of rat apo-S100B(PP) was recalculated using a large n umber of constraints derived from dipolar couplings that were measured in a dilute liquid crystalline phase. The dipolar couplings orient bond vectors relative to a single-axis system, and thereby remove much of the uncertain ty in NOE-based structures. The structure of apo-S100B(PP) indicates a mini mal change in the first, pseudo-EF-hand Ca2+ binding site, but a large reor ientation of helix 3 in the second, classical EF-hand upon Ca2+ binding.