Photosystem Two accessory proteins: structures binding sites and functions

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The Photosystem Two (PSII) complex has quite rightly been described as the 'Engine of Life'. This molecular machine which is found in plants, algae and cyanobacteria is able to use the energy of sunlight to split very stable and abundant water molecules into molecular oxygen, which is released into the atmosphere as a by-product, protons which are used to generate ATP, and 'high energy' electrons which are ultimately used with the ATP to fix atmospheric carbon dioxide into sugar molecules. These carbohydrates can then be used as a fuel and to synthesise biomass for growth. Over the years there has been considerable effort to understand the structure of PSII and how it works. It is now known that PSII is made up of over 20 individual proteins, bound together as a large protein complex in a lipid membrane. Tightly associated with this complex are pigments that harvest the solar energy, small organic molecules that can transport high-energy electrons and a metal cluster comprising one calcium and 4 manganese ions. It is at this metal cluster, buried deep in the protein complex, that water binds and is oxidised to molecular oxygen, giving up its electrons. But PSII is not a perfect machine; it sometimes breaks down, especially when the sunlight is very bright, and has to be repaired. To do this the damaged PSII complex is partially disassembled into a smaller complex, and the damaged protein is recognised, specifically degraded by special proteases found within the membrane, a new protein inserted and the complex reassembled. Without this special repair mechanism PSII would be quickly inactivated in the light and plant growth and oxygen evolution would be inhibited. The purpose of our research is to understand how PSII functions, how PSII is assembled from its component parts and how it is repaired efficiently. Understanding these processes might allow us in the future to enhance photosynthesis in crop plants so that we can increase growth to help satisfy the ever increasing demand for more food and more biomass. This knowledge might also have applications in the design of new, sustainable herbicides or the design of new man-made catalysts that might act as 'artificial leaves' to provide renewable fuels from solar energy. Previous work has identified a number of small proteins that seem to be involved in the assembly, repair or optimal functioning of PSII. We propose to determine the structures of these 'accessory' proteins, so we can see how they are folded in space, and how they bind with PSII. To help do this we have made large amounts of our target proteins in a bacterium, purified the proteins and made crystals which we can use in X-ray diffraction experiments to determine the structure. Alternatively we can also use nuclear magnetic resonance (NMR) spectroscopy which has the advantage that crystals need not be made. We have also been able to make complexes between some of the accessory proteins and either the intact oxygen-evolving PSII complex or smaller protein segments. Analysis of these complexes by x-ray crystallography or NMR will allow us, for the first time, to observe how these accessory proteins engage with PSII at the molecular level. It is important to relate these structural interactions with what is happening in the cell. To do this we have made strains of a cyanobacterium that lack these accessory proteins. By studying PSII assembly and repair and PSII activity in these strains we hope to be able to identify a precise defect in PSII that can be related to the structural results. In this way we will be able learn valuable new information on how the 'Engine of Life' is assembled and maintained in a working state.