Watching Proteins at Work with X-Ray Free Electron Lasers

Proteins are the molecular engines of life. Their broad range of biological tasks and functions is reflected in the large diversity of specific dynamical characteristics they display on a broad time scale. On the femtosecond time scale, atomic bonds vibrate and proteins respond to light stimuli. Amino acid side chains jiggle and wiggle on the pico- to nanoseconds time scale. Within micro- to milliseconds, protein domains change shape and move with respect to each other. Entire pro-teins are synthesized and fold in seconds to minutes.
A large number of experimental techniques exist that each opens a specific window onto mac-romolecular dynamics on a particular time scale. Among those, time-resolved crystallography allows watching proteins at work at atomic resolution. Recently, the advent of an entirely new generation of X-ray sources, so-called X-ray free electron lasers (or “XFEL”), pushed the resolu-tion in time-resolved crystallography down to sub-picoseconds. These devices generate X-ray pulses of very short duration, of the order of a femtosecond (one millionth of a billionth of a sec-ond), with a very high intensity, at the core of an installation several kilometers in length. Exciting possibilities are now opened to structurally follow ultra-fast macromolecular processes such as vision, bioluminescence and other phenomena, which have not been observable to date. As an example, we will show how time-resolved X-ray crystallography at XFELs allowed visualizing photoswitching in fluorescent proteins, which are extensively used as markers in biological imag-ing (see figure).