Detailed snapshots capture photosynthesis in action

One of the most fundamental processes on earth --the photosynthetic splitting of water to generate the oxygen in the air that we all breathe-- is still not fully understood. New X-ray methods have now captured the first detailed images of the machinery that enables this process at room temperature.

HFSP Program Grant holders Uwe Bergmann, Philippe Wernet, Junko Yano and Athina Zouni and colleagues
authored on Mon, 12 December 2016

The living machinery responsible for photosynthesis is still not fully understood. In particular, we don’t know how exactly plants and cyanobacteria harvest energy from sunlight and use it to split water into hydrogen and oxygen, a process that generates the oxygen in the air that we all breathe. New X-ray methods at SLAC National Accelerator Laboratory in Menlo Park, California have captured the first detailed images of the protein complex photosystem II, where this reaction takes places, at room temperature. The research team took diffraction images using the bright, ultrafast X-ray pulses at SLAC’s X-ray free-electron laser – the Linac Coherent Light Source (LCLS). The study is reported in Nature (see reference below).

For many years now, scientists have been trying to understand this process in meticulous detail. Besides its fundamental importance to science, it might help to develop ways to create artificial photosynthesis devices that can serve as potential clean energy sources. The water-splitting reaction involves several steps, each following the absorption of light, and previously only one step, the resting state, had been seen in detail using samples that were frozen.

In this new study, the researchers were able to see two key steps in photosynthetic water splitting under normal conditions of temperature and pressure as it occurs in nature, a big step to decoding how the process works in detail. To do this, the scientists placed droplets of the sample in a solution with small crystallized forms of the large photosynthetic protein complex, photosystem II, on a moving conveyor belt and illuminated the samples with pulses of green light from a laser to initiate the water-splitting reaction. After two light pulses, they captured X-ray diffraction images of the crystals, with a resolution better than 2.5 angstrom.

Schematics of the experimental setup: drops of solution containing photosystem II crystals are deposited on a moving conveyor belt and then flashed with pulses of green laser light. Shortly after that the X-ray laser pulse hits the droplet and the X-rays diffracted from the crystals are recorded for structural analysis. This is repeated several thousand times in order to obtain a full three dimensional reconstruction at high resolution. 

The water-splitting reaction takes place at a tiny metal catalyst within the photosystem II protein, known as the oxygen-evolving complex, which is made up of four manganese atoms and one calcium atom. The complex uses the energy from light to form pure oxygen from two water molecules. The four manganese atoms and maybe other atoms in the catalytic site participate in shuffling electrons through the cycle, but it is unknown where exactly in the complex the relevant water molecules are located or where the oxygen formation occurs. To sort this out, the researchers used ammonia, a water substitute, to narrow down where oxygen atoms from two water molecules combine to form an oxygen molecule. If the ammonia was bound to one of the sites, and the reaction still proceeded, then that site is unlikely to be where the water that will form oxygen binds to the molecule. The results from this study offered a surprise – the data do not seem to support two leading theories for how the reaction proceeds within the oxygen-evolving complex.

In future studies, the researchers hope to capture more images at different steps of the process, which will allow them to further refine the details of the water-splitting reaction. In fact, the chemistry of photosynthetic water-splitting is very unusual and learning exactly how this process works will not just be a breakthrough in our understanding, it can help in the development of solar fuels and renewable energy.

The international research team is a long-standing collaboration between Berkeley Lab and SLAC and includes Humboldt University and Helmholtz Zentrum Berlin in Germany, Umeå University and Uppsala University in Sweden, Stanford University, and the Brookhaven National Laboratory in the USA and the University of Oxford in the UK. Although this is a very large collaboration with many funding sources, the HFSP funding was especially important as it supported personnel and the work of all four PIs on the grant, critical to this publication.

Reference

Structure of photosystem II and substrate binding at room temperature. Iris D. Young, Mohamed Ibrahim, Ruchira Chatterjee, Sheraz Gul, Franklin D. Fuller, Sergey Koroidov, Aaron S. Brewster, Rosalie Tran, Roberto Alonso-Mori, Thomas Kroll, Tara Michels-Clark, Hartawan Laksmono, Raymond G. Sierra, Claudiu A. Stan, Rana Hussein, Miao Zhang, Lacey Douthit, Markus Kubin, Casper de Lichtenberg, Long Vo Pham, Håkan Nilsson, Mun Hon Cheah, Dmitriy Shevela, Claudio Saracini, Mackenzie A. Bean, Ina Seuffert, Dimosthenis Sokaras, Tsu-Chien Weng, Ernest Pastor, Clemens Weninger, Thomas Fransson, Louise Lassalle, Philipp Bräuer, Pierre Aller, Peter T. Docker, Babak Andi, Allen M. Orville, James M. Glownia, Silke Nelson, Marcin Sikorski, Diling Zhu, Mark S. Hunter, Thomas J. Lane, Andy Aquila, Jason E. Koglin, Joseph Robinson, Mengning Liang, Sébastien Boutet, Artem Y. Lyubimov, Monarin Uervirojnangkoorn, Nigel W. Moriarty, Dorothee Liebschner, Pavel V. Afonine, David G. Waterman, Gwyndaf Evans, Philippe Wernet, Holger Dobbek, William I. Weis, Axel T. Brunger, Petrus H. Zwart, Paul D. Adams, Athina Zouni, Johannes Messinger, Uwe Bergmann, Nicholas K. Sauter, Jan Kern, Vittal K. Yachandra & Junko Yano. Nature (2016) doi:10.1038/nature20161.

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