Raman Spectroscopy as a tool for probing protein structure

Principally through the efforts of crystallographers, we are being presented with an ever expanding molecular view of the biological world. Although this brings into focus many questions regarding the mysteries of function, techniques are need that facilitate the transition in our understanding from structure to function. Raman spectroscopy is one of these. Raman spectroscopy involves analysing the scattered photons from a laser beam focused into the sample. The inelastic scattered photons provide information on molecular vibrations that, in turn, yield data on molecular conformation and environment. Thus, Raman spectroscopy can provide very detailed information on little sites in a much larger macromolecules complex. Raman spectroscopy has become a versatile tool in protein science and biotechnology thanks to the improved instrument sensitivity which has increased the signal-to-noise ratio. Thus, this technique can be successfully used for determination of protein secondary structure, identification of metal coordination sides, hydrogen bonding, oxidation state of cysteine residues, local environments of aromatic residues (i.e. Tyr, Trp), protein-ligand and -DNA interactions, etc. For these reasons it has been used to prove or refuse models (i.e. in enzyme or receptor mechanisms), to improve models, and to suggest mechanisms, since the advantage of this spectroscopic technique is its extreme sensitivity to changes in structure and molecular interaction. In particular, in our lab, Raman spectroscopy has been recently used for evaluating the changes in the protein structure resulting from radical exposure and the susceptibility of some sites to radical attack.