Environmental constraints in the study of flexible segments of proteins.

The structural problem posed by ill-defined segments in protein structures is similar to those encountered in the study of most peptide hormones, with terminal tracts resembling linear peptides and loops resembling cyclic peptides. The conformational preferences of short linear peptides in solution can be influenced by the use of solvent mixtures of viscosity higher than that of pure water but comparable to that of cytoplasm. In order to check whether it is possible to use these media in the structural study of proteins, we undertook an exploratory study on BPTI in a mixture of dimethylsulfoxide and water. The complete assignment of BPTI in an 80:20 (by volume) DMSO-d 6/water cryomixture at two temperatures showed that all resonances parallel those in water, hinting at the persistence of the correct protein architecture, which is also confirmed by NOESY experiments. In addition to the NOEs present in the aqueous solution it was possible to detect numerous new cross peaks, in particular from residues belonging to the less-defined regions. The new cross peaks do not originate from spin diffusion and are consistent with the best NMR structure and with the X-ray structures of BPTI.