This project is divided in five workpackages and a methodological subproject that focuses on elucidating how different functional modules in complex I establish a 300 Å charge transport process. The project resulted in 30 peer-reviewed publications, and several manuscripts that are in review or preparation during the finalizing of the project. Many of the publications are published in high-impact journal (JACS, PNAS, Science Adv, Nature Comms) supporting that our project team has secured a high level of research. The work has been presented in multiple leading conferences in the field and communicated in review articles to reach both experts and non-expert audiences. A brief summary of each workpackage (WP) is given below.
In WP1, we have studied functional elements that are used for conversion of chemical energy to redox energy. We have studied how electrons enter the complex I with the redox cofactor NADH, and how this module further bifurcates the electrons into a chain of FeS redox-cofactors. We identified protein residues that are responsible for the catalytic activity and how certain point mutations may enhance the production of reactive oxygen species (ROS), linked to the development of mitochondrial disease. In WP2 and WP3, we have studied functional elements that mediate long-range electron transfer and establish the catalytic core of complex I. We have identified electronic and structural changes linked to the electron transfer process as well as its energetics along the FeS chain. Moreover, we have established the function of key elements that enables complex I to reduce the quinone substrates, and conformational changes linked to this process. In WP4, we have studied elements mediating redox energy for proton pumping, and identified a novel substrate binding-site that may activate the proton pumping activity of the enzyme. In WP5, we have studied how different protein modules establishing the long-range proton-pumping machinery, and how the biological membrane surroundings modulate the biological activity. During the project, we have developed computational simulations methods, linked with protein design and biophysical experiments, that has enabled us to probe the structure and function of the catalytic machinery in complex I on a broad range of time-scales and spatial resolutions.
During the project, we identified functional elements of the Complex I machinery that are responsible for the long-range proton transport process (PNAS 118:e2019498118; PNAS 114:E6314; JACS 142:13718; JACS 142:21758). Based on these, we suggested key principles by which Complex I enables the long-range energy transduction process by coupled conformational and hydration changes, triggering protonation reactions that propagate across the complete (>200 Å) membrane domain (PNAS 114:E6314; JACS 142:13718). We could also resolve how the active site of Complex I reduces the quinone substrates (PNAS 112:11571; PNAS 114:12737; JACS 139:16282), how the enzyme oxidizes biological electron carriers such as NADH (JACS 141:5710), we identified novel substrate binding sites (PNAS 115:E8413; Nat Commun 11:5261), and determined how the lipid environment affects its functional dynamics (Science Adv 5:eaav1850). Moreover, our work unraveled how modular adaptations of the Complex I architecture lead to new functionality that support photosynthetic energy conversion, including the concentration of CO2 in cyanobacteria (Nat Commun 11:494), production of hydrogen gas (H2) in archaea (JACS 143:20873), as well as the electron bifurcation in acetogenic bacteria living in extreme conditions (JACS 145:5696). Despite significant structural differences to other energy transducing enzymes, our studies also allude to conserved mechanistic principles that could be generally employed in biological energy conversion (Acc Chem Res 54:4462). The integrated computational-experimental approaches that we developed to address these mechanistic questions, allowed us to further design artificial proteins as frameworks for testing biological hypotheses (Nat Commun 12:1895), without the "evolutionary baggage" of natural proteins. These methods will be used in future work to address challenging mechanistic questions in biology. The ERC project resulted in several articles published in top journals (Nature Comm, PNAS, Sci. Adv., JACS). The work was disseminated in public talks in conferences, invited talks at various universities, as well as to the general public.
