Serial Femtosecond Crystallography Opens a New Era in Membrane Protein Structural Biology

Abstract

Serial Femtosecond Crystallography (SFX) provides a novel concept for structure determination, where X‐ray diffraction “snapshots” are collected from a fully hydrated stream of nanocrystals, using femtosecond pulses from the world's first high energy X‐ray free‐electron laser, the Linac Coherent Light Source. The large membrane protein complex Photosystem I was used as the model system for the first serial femtosecond crystallography experiments that showed the proof of concept that diffraction of nanocrystals can be observed using femtosecond pulses that are 1012 stronger than 3rd generation synchrotron sources and destroy any material that is placed in its focus [1],[2]. By using femtosecond pulses briefer than the time‐scale of most damage processes, femtosecond nanocrystallography overcomes the problem of X‐ray damage in crystallography [3] and extends to atomic resolution [4],[5]. It is also applied to membrane proteins crystallized in lipidic environments [6],[7], and has recently led to the structure determination of the Angiotensin receptor which regulates the blood sugar level and is one of the major targets for drugs fighting hypertension [8] and the structure of the rhodopsin‐arrestin complex [9]. Femtosecond crystallography also opens a new avenue for determination of protein dynamics. First experiments on the proof of principle for time resolved serial femtosecond nanocrystallography have been performed on Photosystem I‐ferredoxin [10] and Photosystem II nano‐crystals [11] and conformational changes of the Mn4Ca cluster and its protein environment were observed for the first time in the transition from the dark to the double excited state [12]. Very recently we were able to show that TR‐SFX studies extend to atomic resolution using the photoactive yellow protein as a model system [13]. This pioneering work paves the way for the determination of molecular movies of the dynamics of membrane proteins “at work” in the future including the determination of molecular movies of water splitting.