Domain orientation and dynamics in multidomain proteins from residual dipolar couplings.

The data most commonly available for the determination of macromolecular structures in solution are NOE based distance estimates and spin-spin coupling constant based dihedral angle estimates. This information is, unfortunately, inherently short-range in nature. Thus, for many multidomain proteins, little information is available to accurately position weakly interacting domains with respect to each other. Recent studies of proteins aligned in dilute liquid crystalline solvents have shown the utility of measuring anisotropic spin interactions, such as residual dipolar couplings, to obtain unique long-range structural information. In this work, the latter approach is taken to explore the relative domain orientation in a two-domain fragment from the protein barley lectin. An approach based on singular value decomposition as opposed to simulated annealing is used to directly determine order tensors for each domain from residual (15)N-(1)H dipolar couplings, and the limitations of the two approaches are discussed. Comparison of the order tensor principal axis frames as separately determined for each domain indicates that the two domains are not oriented as in the crystal structure of wheat germ agglutinin, a highly homologous protein ( approximately 95% sequence identical). Furthermore, differences in the order tensor values suggest that the two domains are not statically positioned but are experiencing different reorientational dynamics and, to a large degree, may be considered to reorient independently. Data are also presented that suggest that a specific association occurs between one domain and the lipid bicelles comprising the liquid crystal solvent.