Calculation of ensembles of structures representing the unfolded state of an SH3 domain

The N-terminal SH3 domain of drk (drkN SH3 domain) exists in equilibrium between a folded (F(exch)) and an unfolded (U(exch)) form under non-denaturing conditions. In order to further our previous descriptions of the U(exch) state, we have developed a protocol for calculating ensembles of structures, based on experimental spectroscopic data, which broadly represent the unfolded state. A large number of unfolding trajectories were generated, starting from the folded state structure of the protein, in order to provide a reasonable sampling of the conformational space accessible to this sequence. Unfolded state ensembles have been "calculated" using a newly developed program ENSEMBLE, which optimizes the population weights assigned to each structure based on experimental properties of the U(exch) state. Pseudo-energy terms for nuclear Overhauser effects, J-coupling constants, (13)C chemical shifts, translational diffusion coefficients and tryptophan ring burial based on NMR and fluorescence data have been implemented. The population weight assignment procedure was performed for different starting ensembles. Small numbers of structures (