Task 3 : Supramolecular organisation of membranes and membrane proteins

(PARTNERS 1, 2, 4, 5, 6, 7)

Quinones are key players of the electron transfer chain, diffusing easily in the membrane and playing an important role in electron transfer. The synthesis of ubiquinone is still poorly established, although it is highly conserved within all living organisms and a ubiquinone deficit in humans results in severe mitochondrial disorders and pathologies. Using a bacterial model, Partner 6 identified a large supramolecular complex (Ubi complex) responsible for ubiquinone biosynthesis and several new key protein partners (UbiK and UbiJ) (Loiseau, J Biol Chem, 2017). Intriguingly, the complex commutes between the membrane and the cytosol, in a process seemingly controlled by lipid binding.

The control of energy input is crucial, especially in photosynthesis where significant fluctuations in light intensity occur. Cells need to harvest sufficient energy without overloading the ETC.  An important solution to this problem is the redistribution of light harvesting complexes between supercomplexes. Partners 1, 4 and 5 have made several breakthroughs on the regulation of the redistribution of light between photosystems I and II. The light harvesting complexes (LHCs) are able to redirect light energy towards one or other of the photosytems depending on need, or to dissipate energy in case of stress (Fig. 6). We showed that microalgae redistribute their energy rather than dissipating it as previously thought, thereby allowing a well-balanced influx of excitation energy (Nawrocki, Nat Plants, 2016). We further showed (in collaboration with J. Alric, CEA Cadarache) that the Stt7 kinase controls the redistribution of LHCII bound to cytochrome b6f, not far from the quinone reducing site Qi. This allowed us to propose a new mechanism of how light flow is regulated through the redox poise; furthermore, it explains how the signal is transmitted through the membrane (Dumas, Proc Natl Acad Sci U S A, 2017). Lastly, we provided clear evidence for a regulatory circuit that controls LHCII translation in response to light quantity in unicellular green algae (Berger, Plant Physiol, 2016).

In photosynthesis, efficient conversion of carbon dioxide into organic matter requires a tight control of the ATP/NADPH ratio. Partner 1 has shown (through national and internal collaborations) that diatoms regulate ATP/NADPH through extensive energetic exchanges between cp and mt. This interaction comprises the re-routing of reducing power generated in the cp towards mt and the import of mt ATP into the cp, and is mandatory for optimized carbon fixation and growth. This process likely contributed to the proliferation of diatoms, one of the most ecologically successful classes of marine eukaryotes in contemporary oceans. Over the past 30 million years, they have helped to moderate the Earth’s climate by absorbing CO2 from the atmosphere, sequestering it via the biological carbon pump and ultimately burying organic carbon in the lithosphere. The proportion of planetary primary production by diatoms in the modern oceans is similar to that of terrestrial rainforests (Bailleul, Nature, 2015). The recruitment of Benjamin Bailleul to the lab of Partner 1 has brought an exceptional biophysical expertise in coupling electron transfer with physiological mechanisms to the Dynamo consortium.

In the ETC, the insertion of a loop called the Q-cycle increases the yield of translocated protons per transferred electron. This is done at the cost of introducing an electron-bifurcation step in the reaction that creates a rate-limiting step and a potential site for reactive oxygen species (ROS) formation in the Qo quinone oxidizing site of the complex. Most advances in the past came from studies on the mitochondrial complex bc1 or its related proteobacterial counterpart. Mechanisms are still controversial, especially that concerning the bifurcation step. Partner 5 has therefore chosen to transpose these questions on homologous complexes that uses other quinones: plastoquinone (in photosynthesis) or menaquinone (in the respiration of Gram-positive bacteria). We isolated supercomplexes of the thermophilic bacterium Geobacillus stearothermophilus. We presented preliminary results in the mid-term report and we have now completed its full redox characterisation (Fig. 7A) (Bergdoll, Biochim Biophys Acta, 2016). We observed that redox groups have a mid-point potential much lower than previously expected. Our analysis validates older models of Q-cycles, discussed in a recent review on biological bifurcation.

DYNAMO has fostered a collaboration between Partners 1, 5 and 7 to build a photosynthetic bio-photovoltaic cell. The above-mentioned Q-cycle, while increasing the overall yield, does so at the cost of introducing a rate-limiting step. It follows that it should in principle be possible to divert part of the electron flow upstream of the cytochrome b6f at the level of photosystem II without impeding the overall photosynthetic process. Furthermore, a careful design could allow harvesting of current from a living self-replicating electrochemical solar cell. To do so, we first tested several quinones as new electron carriers to transfer current from the algal cp to the collecting electrode and selected those that did not impede the electron flow necessary for photosynthesis (i.e. not binding to the QB site). Second, the accessibility of the QA electron transfer site was increased by mutagenesis, thus creating an electron extraction site in photosystem II. Third, electronic currents were harvested in an electrochemical cell (Fig. 7B) (Fu, Nat Commun, 2017; Longatte, Chemical Science, 2018). This approach was tested successfully using other quinones (Longatte, Photochem Photobiol Sci, 2016) and provides an important proof of concept for future design of more efficient bio-photovoltaic cells.

Partner 6 has explored different bio-inspired chemical strategies to store solar energy as chemical energy (fuels). Energy storage can be achieved via water splitting or carbon dioxide reduction. For this purpose, we developed a variety of new bio-inspired molecular or solid catalysts, mimicking active sites of metallo-enzymes, for water oxidation, proton reduction and CO2 reduction with remarkable performance (Elgrishi, Chem Soc Rev, 2017). In addition, an artificial enzymology approach has led to original hybrid systems, combining a protein host and a synthetic molecular catalyst, which behave as artificial hydrogenases, catalyzing water conversion to hydrogen (Porcher, Angew Chem Int Ed Engl, 2015).

In their ensemble, the studies we performed in the 3 tasks replied very well to the goals we set ourselves for DYNAMO and have laid the foundation for us to build on this progress, to expand our future fundamental research efforts to fuel the development of additional innovative tools that can be adapted to tomorrow’s energy needs.