14-PSIL MAGIC: a multi-tiered approach to gaining increased carbon

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We are building on progress that includes expression of pHR in E. coli, cyanobacteria and plants. These advances have put us in a strong position to deliver within the next few years. Mathematical modeling has validated the idea of using a light-driven ion pump for concentrating CO2; it now remains to assemble and express these pumps and validate function in chloroplasts. The idea to use scaffolds to concentrate CO2 at RuBisCO remains a goal, but our strategies have changed in light of new understanding of the interplay between diffusion and kinetics. We have successfully expressed scaffold proteins in cyanobacteria and plants, demonstrating that they can be both targeted to specific sites and that they function to recruit their respective substrates. Our mathematical models predict that the original idea of utilizing these constructs to enhance channeling of CO2 to RuBisCO will have negligible impact on CO2 assimilation. We need now to confirm this prediction in our cyanobacterial systems. A rethinking of the problem of concentrating CO2 at RuBisCO in C3 plants leads to development of a new approach. Our mathematical models highlight the poor CO2 capture probability of RuBisCO as a major constraint. Here, we propose designs to slow the diffusion rate of CO2 in the stroma and increase assimilation by introducing transient (stationary) binding sites near RuBisCO (a CO2 'sponge'), effectively enhancing the native characteristics recently identified in photosynthetic systems. We will use the cyanobacterial system to screen and optimize this approach and will use the scaffolds now proven in our hands to translate these to chloroplasts. Finally, we previously lacked the ability to quantify performance, ie. HCO3- concentration gains. This capability is now available through a lipid vesicle technique.

This proposal is for fundamental research to develop new conceptual approaches relevant to ideas emerging within the international plant, systems and synthetic biology communities. The research will stimulate thinking around strategies for modelling and for applications of synthetic biology in plants, especially in relation to photosynthesis, and it should strengthen methodologies relevant at many levels from cell to crop engineering. Thus, the research is expected to benefit fundamental researchers and, in the longer-term agriculture and industry, through conceptual developments and approaches to improving carbon capture by plants. The research will feed into higher education training programmes through capacity building at the postgraduate and postdoctoral levels. Additional impact is proposed through public displays and the development of teaching resources building on the background work for this proposal. Finally the research will help guide future efforts in applications to agricultural/industrial systems. The applicants have established links with industrial/technology transfer partners and research institutes to take advantage of these developments. Further details of these, and additional impacts will be found in Part 1 of the Case for Support and in the attached Impact Pathways.