Decreasing the oxygenase activity of Rubisco: a synthetic biology approach

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Photosynthesis is the process whereby plants convert atmospheric carbon dioxide to organic matter (biomass) using energy from sunlight. Photosynthesis powers plant growth, crop productivity and all life on earth. The first step in photosynthesis combines carbon dioxide with a sugar containing 5 carbon atoms to make two sugar molecules each containing 3 carbon (C3) atoms in a process called carbon dioxide fixation. The C3 sugars then combine to regenerate the 5 carbon accceptor and also to form glucose. Glucose is used as the feedstock to synthesise all other molecules in the plant. Chemical reactions in plants, such as the first step in photosynthesis, only happen quickly enough if they are speeded up by proteins known as enzymes. Rubisco is the enzyme that speeds up carbon dioxide fixation. Unfortunately this enzyme has built-in inefficiencies. It is slow, so plants need to spend energy producing it in large quantities. It also has a side reaction in which oxygen competes with carbon dioxide to react with the 5C acceptor. The consequence of this side reaction (known as the oxygenase reaction) is that less C3 product is formed and photosynthesis rate is slower than it could be. Some algae and plants have developed elaborate mechanisms to increase the concentration of carbon dioxide near Rubisco, thereby decreasing oxygenase activity and increasing the efficiency of photosynthesis. Unfortunately, many of our major crops (e.g. rice, wheat, potatoes and pulses) do not have this mechanism and various approaches to reducing their oxygenase activity are being intensively investigated. To contribute to this effort, we are proposing a pilot study to assess the potential of physically linking Rubisco to another enzyme (carbonic anhydrase, CA) which, under the right conditions, could deliver a high concentration of carbon dioxide to Rubisco and reduce the wasteful oxygenase activity. To achieve this aim we propose to reconfigure the carbon dioxide fixing step of photosynthesis by engineering the bacterium Synechocystis. We will prepare a synthetic gene which, when introduced into Synechocystis will cause it to produce a protein scaffold that can bind both Rubisco and CA. The modified strain will also contain forms of Rubisco and CA that have been engineered with tags that allow them to bind to the scaffold protein. This engineered organism will allow us to test the proposal that close proximity of Rubisco and CA increases the efficiency of photosynthesis by decreasing the "wasteful" oxygenase activity.