Disordered Regions and Protein Crystallography

Structure-based drug design is one of the most powerful strategies for current drug discovery. While this strategy requires a high-resolution structure of a target protein, most drug target proteins are not readily crystallizable. Many generally well structured proteins have disordered regions [50] that may inhibit crystallization and/or cause poor diffraction. The identification of disordered regions can help in designing new constructs that retain structure and biological function, but are depleted of disordered regions. While a number of approaches to identifying disordered regions are available, ranging from computational stability calculation to limited proteolysis, none of them have provided reliable identification of disordered regions at the pace required.

Since the exchange rates of backbone amide hydrogens are most influenced by their involvement in hydrogen bonding to other portions of the protein, measurement of exchange rates allows direct identification/localization of ordered/ disordered regions in the protein. Amide hydrogens in disordered regions exchange very rapidly because the hydrogen bonds to amides in such regions are predominantly with solvent water most of the time. We have recently shown that suitably performed DXMS studies can rapidly localize such disordered regions, and guide the design of well crystallizing constructs selectively depleted of disorder [44].

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