Theoretical study of the cytochrome bc1 complex reaction mechanism

The bc1 complex is an enzyme which plays a critical role in energy production through photosynthesis and cellular respiration. Its biochemical function relies on the so-called Q- cycle, which is well established and operates via quinol substrates that bind the protein at their active sites. Despite decades of research, the quinol-protein interaction that initiates the Q-cycle has not yet been completely described. Furthermore, the initial charge transfer reactions that take place following quinol binding, lack a physical description. The present dissertation presents a comprehensive study of the primary reaction mechanism of the bc1 complex from the photosynthetic purple bacterium Rhodobacter capsulatus. By using theoretical methods, such as classical molecular dynamics simulations and quantum density functional theory calculations, we investigated the molecular structure and function of the bc1 complex to provide a quantitative description of the primary events that occur during and after quinol binding. First, we studied the binding motifs of a quinol molecule at its active site of the bc1 complex. Our investigations suggested a novel configuration of amino acid residues responsible for quinol binding and provided new insights into the role of the different amino acid residues that hold the quinol molecule at the Qo-site of the bc1 complex. The calculations were performed for two Qo-site models, differing in the protonation state of a histidine residue that is proven to be fundamental in the binding and further reaction. The findings were consistent with some features of earlier molecular dynamics simulations, but these identified also rearrangements of binding site residues not discussed previously. Secondly, by exploring all possible single and paired-charge reaction pathways, we studied the electron and proton transfer reactions that trigger the Q-cycle. In particular, the coupled nature of the first electron and proton transfer reactions was revealed, accompanied by a transition path that connects the configurations of the Qo-site prior to and after the charge transfers. All the calculations were performed for the two Qo-site models; however, the protonated-histidine model was found to be more favorable for the reactions. Key structural elements of the bc1 complex that trigger the charge transfer reactions were established, demonstrating the importance of the environment in the reaction, which is furthermore evidenced by free energy calculations of the reaction. Once the primary charge transfer was identified as a proton-coupled electron transfer (PCET) reaction, we focused our efforts on obtaining the reaction rate constant. For this purpose, we established the adiabaticity of the PCET reaction, as well as other parameters. These included the vibronic couplings (electronic coupling and overlap of the proton vibrational states at the reactant and product states), and the dependence of the rate constant on the proton donor-acceptor distance. Finally, we calculated the kinetic isotope effect of the PCET reaction. Based on the obtained values, and by comparing with previous experimental investigations, we found strong indication that the primary PCET reaction is indeed the rate-limiting step of the Q-cycle.Ope