# Artificial Photosynthesis: Is Computation Ready for the Challenge Ahead?
## 3 Protein ET Mechanisms
Nature favors proteins in which the average distance between redox centers is within 1.4 nm, for which the ET is robust via a tunneling mechanism, while for longer distances the ET flow is weaker and via a less effective hopping mechanism [121]. As the distance between active sites cannot be always rearranged, nature found the solution to shift the ET process from one-step tunneling to multi-steps hopping mechanism [124; 125]. Tunneling through protein is mediated via amino acid residues, which merely act as conductive elements; on the other hand, during hopping some amino acids are reversibly reduced/oxidized and serve as intermediate states in between the donor and acceptor. As a result, if the intermediate state is reduced by the donor, consequently reducing the acceptor too, there is electron hopping. Conversely, if the intermediate state is oxidized by the acceptor leading to oxidization of the donor, there is hole hopping. Adding several intermediate states strongly decrease the donor-acceptor distance, replacing the long, slow tunneling into a cascade of short and fast ET hopping steps. ET in Small Proteins  
One of the most studied model system for ET mechanism is azurin (Figure 10). Despite not being a LHC, it is often used as model to compute the ET ability either in solvent or on surface, thus being a source of improvement for different computational methods. It is a small blue copper protein and serves as electron shuttles to mediate different ET reactions in biological metabolisms [119; 126; 127]. In order to assess the ET mechanism of any protein containing a redox center, its reduction potential should be computed with high accuracy, as is one of the key pa rameter needed for the ET. The computational determination of individual reduction potentials are affected by large systematic errors with respect to experiments, which make this task difficult to perform. Main components contributing to the reduction potential include electrostatics [128; 129; 130], metal-ligand interactions [131], hydrogen bonding interactions [132] and the constrain of the protein [133; 134]. Common methods to compute the reduction potential consider MD simulations [135], continuum electrostatics methods [136], DFT [137; 138], as well as hybrid QM/MM methods [139]. Recently, a comprehensive QM/MM approach for a large dataset of reduction potentials of mutants and an efficient and reliable computationally procedure has been reported [140]. The authors consider both continuum electrostatics and DFT methods on 24 azurin mutants, and found that the first gives good agreement with experimental observation despite its low computational cost and somehow high degree of approximations. Yet, it failed in properly describing the reduction potentials when explicitly hydrogen bonds are present and when hydrophobic axial ligands are considered. When hydrogen bonds are created/deleted through mutations, DFT or QM/MM approaches are needed. The introduction of hydrophobic ligand lead to a failure of DFT in predicting reduction potentials, and the authors suggest the introduction of a \"hydrophobicity factor\" through the Kyte-Doolittle hydrophobicity index \(\Delta\)KD [141]. In addition, when a combination of hydrogen bonds and hydrophobic axial ligands are present, DFT should be used to compute the reduction potential of the redox center (primary sphere) and the continuum electrostatic model for changes in the secondary sphere and the resulting total reduction potential is the sum of the two terms, in an approach similar to the ONIOM method. Finally, the authors reported an extensive prediction study on over 124 azurin mutants, for which they also provide a general scheme to be followed, in an elegant way, which might be used also for different PPCs (Figure 11). As already discussed in the presented review, in order to accurately determine the redox properties of metalloprotein, both the electronic structure of the redox center and the environment must be considered.