Synchrotron-based Single-Shot Infrared Spectroscopy at BESSY II

Proteins are among the key players in the cells of living organisms, and one of the most important techniques to study their structure and function is infrared (IR) spectroscopy. State-of-the-art time- resolved IR spectroscopy methods employed to follow protein kinetics at a timescale of milliseconds and below are often limited to cyclic systems in which exactly the same reaction can be triggered in quick succession and multiple times, without any substrate loss(1). However, the majority of systems in nature is either not cyclic or has a long cycling time, making time-resolved IR measurements either impossible or very demanding. For example, vertebrate rhodopsin, the protein that initiates vision in our eyes, undergoes an irreversible activation cascade after light absorption. Although this protein is in the focus of scientific interest in pharmacology and medicine because it constitutes the prototype of a huge family of protein receptors mediating basic functions of life such as vision, smell, taste and drug reception(2), no time-resolved IR spectra of its activation cascade have been published so far. We developed a mid-IR spectrometer with microsecond time resolution specifically tailored for such irreversible processes. It uses the brilliant synchrotron radiation source of BESSY II(3) in combination with a fast linear detector array in a unique diffraction limited dispersive spectrometer concept(4).
Here we present the first results using this novel spectrometer on several light-activated protein systems, amongst others, vertebrate rhodopsin.

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2. Hilger, D., Masureel, M. & Kobilka, BK., Structure and dynamics of GPCR signaling complexes. Nat. Struct. Mol. Biol. 25, 4–12 (2018)
3. Peatman WB. & Schade U., A brilliant infrared light source at BESSY. Rev Sci Instrum. 72, 1620 (2001).
4. Schade U., Ritter, E., Hegemann, P., Aziz, EF. & Hofmann, KP., Concept for a single-shot mid-infrared spectrometer using synchrotron radiation. Vib. Spectrosc. 75, 190 (2014).