Raman-assisted crystallography of biomolecules at the synchrotron: Instrumentation, methods and applications

Raman spectroscopy is a powerful technique that, in recent years, has been successfully coupled to X-ray crystallography for the analysis of biological macromolecular systems. The complementarity between both techniques is illustrated at multiple stages, including sample preparation, data collection and structural interpretation with a mechanistic perspective. The current state of instrumentation is described, focusing on synchrotron based setups. Present and future applications of Raman microspectrophotometry are reviewed with reference to recent examples dealing with metallo-, photosensitive-, and redox-proteins. The added value of Raman microspectrophotometry to assess X-radiation damage is discussed, and its applicability to investigate crystalline DNA molecules is also emphasized. This article is part of a Special Issue entitled: Protein Structure and Function in the Crystalline State.

Research Highlights
► Raman spectroscopy is an emerging technique at synchrotrons.
► Raman microspectrophotometry provides information highly complementary to macromolecular crystallography, shedding light on the mechanism of enzymes.
► Raman microspectrophotometry is a valuable non-invasive tool for quantifying radiation damage in macromolecular crystallography and providing the zero-dose point.