The vast majority of biomolecular processes are controlled or facilitated by water interactions. In enzymes, regulatory proteins, membrane-bound receptors and ion-channels, water bound to functionally important residues creates hydrogen-bonding networks that underlie the mechanism of action of the macromolecule. High-resolution X-ray structures are often difficult to obtain with many of these classes of proteins because sample conditions, such as the necessity of detergents, often impede crystallization. Other biophysical techniques such as neutron scattering, nuclear magnetic resonance and Fourier transform infrared spectroscopy are useful for studying internal water, though each has its own advantages and drawbacks, and often a hybrid approach is required to address important biological problems associated with protein–water interactions. One major area requiring more investigation is the study of bound water molecules which reside in cavities and channels and which are often involved in both the structural and functional aspects of receptor, transporter and ion channel proteins. In recent years, significant progress has been made in synchrotron-based radiolytic labeling and mass spectroscopy techniques for both the identification of bound waters and for characterizing the role of water in protein conformational changes at a high degree of spatial and temporal resolution. Here the latest developments and future capabilities of this method for investigating water–protein interactions and its synergy with other synchrotron-based methods are discussed.
Homogeneously N-glycosylated proteins derived from the GlycoDelete HEK293 cell line enable diffraction-quality crystallogenesis