The construction of light-harvesting electrodes based on the integration and exploitation of components from the natural photosynthesis is an emerging field in current research and potential can be seen to contribute to the development of a bio-based economy. In this thesis the focus was applied to produce photobioelectrodes by the utilization of photosystem I, as one of the major light-to-charge separating super-complexes from the thylakoid membrane in the oxygenic photosynthesis. The electrical connection of this complex with an inorganic electrode material is often rather difficult. Hence, a new strategy has been investigated by coupling PSI by means of the small redox protein from the respiratory chain cyt c to the electrode. The essential features of the biohybrid photoelectrodes will be summarized hereinafter.
In the first part of this thesis (P4.1), the direct interaction between both biocomponents has been addressed, whereas the experimental conditions for the assembly on a SAM modified gold electrode have been revealed. Here, PSI was able to assemble in a densely packed layer on a cyt c monolayer under buffer conditions of low ionic strength and at a neutral pH. It has further found, that cyt c promotes the binding of PSI to the surface. The photobioelectrode exhibited a photocurrent of about 1 µA cm-2 at -0.1 V vs. Ag|AgCl. The direction of the photocurrent is solely cathodic, since at higher potentials (+0.5 V vs. Ag|AgCl) only very small anodic currents have been observed. The light-induced catalysis started at the redox potential of cyt c. This demonstrates, that the electrons are first transferred from the electrode to cyt c further reducing the PSI in a second step. It also shows, that PSI can be assembled on cyt c preferentially with the luminal side down The study also reveals, that multiple alternately assembled layers of cyt c and PSI can be built up with an additional contribution to the overall detected photocurrent.
The second part of this thesis (P4.2) has aimed for the further improvement of the photobioelectrode and for the elucidation of a deeper understanding of the connection of PSI with cyt c. It was found that cyt c and PSI were able to produce artificial complexes already in solution. Both biocomponents can be deposited as a thick layer from a mixed solution on a modified gold surface. The deposited amount strongly depends on the molecular ratio of cyt c to PSI and on the assembling time. The surface concentration of electrically addressable cyt c
Summary surface was limited and DNA, as a polyelectrolyte structural scaffold, was introduced to circumvent the saturation of biomolecules on the electrode. With a layer-by-layer approach, the photocurrent was greatly enhanced up to 27 µA cm-2 for an 8 bilayer electrode. In addition the long-term stability of the photobioelectrode was improved (65 % activity after 1 month’s storage). The photophysical parameters of the cyt c / PSI electrode revealed high quantum efficiencies of 4.9 % (EQE) and 27 % (IQE), with a moderate turnover number of PSI (21 e- PSI-1 s-1).
In the third part of this thesis (P4.3) mesoporous indium tin oxide electrodes have been developed for the integration of cyt c and PSI into a 3D transparent electrode. The construction process of µITO electrodes has been investigated in detail with respect to structural scalability. It was possible to integrate the large membrane super-complex PSI and the small redox protein cyt c with monolayer coverage with respect to the electrochemically active surface area of the µITO electrode. The produced photocurrents reached up to 150 µA cm-2 (for a 40 µm thick or a 6x layer electrode) and the improvement of the heterogeneous electron transfer constant, ks of cyt c to the electrode has been identified to play a crucial role in this system. The internal quantum efficiency of this photobioelectrode achieved a very high value of 39 %, which is extraordinary high if compared with other strategies using PSI. A further advantage of this biohybrid electrode can be seen in the ease of preparation. The 3D electrode can be produced using a spin-coating procedure within hours, while the integration of the protein components takes only several minutes.
From all studies performed within this thesis it can be concluded, that the interaction and communication of cyt c with PSI plays a crucial role. The binding of both biocomponents seems to be of non-specific mainly electrostatic nature. Computational analysis of putative contact sites shows, that cyt c is able to bind to different parts of PSI, including the luminal and stromal side, as well as hydrophobic regions in the membrane section of the protein complex. This feature explains, why both biocomponents can be assembled onto each other and form co-complexes in solution, whereby a protein multilayer or co-assembly are feasible.
However a preferential binding to the luminal side can also be shown and explains the docking of PSI to a cyt c monolayer. In the multilayer system PSI molecules far away from the electrode surface can be functionally connected, and the generated photocurrent exhibits a linear relationship with the protein amount deposited. This can be explained by the efficient self-exchange between the cyt c network surrounding PSI.
Summary The comparison to other photobioelectrode strategies using PSI displays a remarkable development over the studies provided by this thesis. In terms of performance, quantum or molecular efficiency, the cyt c / PSI photobioelectrodes P4.2, but especially P4.3 belong to the best systems to date, making them candidates for the generation of more complex signal chains. Especially the light-to-current conversion efficiency of P4.3, expressed by the internal quantum efficiency can reach up to 39 %, while the molecular efficiency expressed by the turnover number can be as high as 35 e- PSI-1 s-1. Furthermore, the systems even provide a high performance under lower light conditions, which once again shows that efficient signal pathways have been created.
Taken together, these three studies demonstrate the potential and limitations of constructing a photobiohybrid electrode based on PSI, when cyt c was used for the wiring to the electrode.
Within this thesis, the working conditions of the cyt c/PSI system have been revealed, while optimization of performance have been done by the choice of electrode material and protein arrangement, e. g. by using DNA. The photo-induced signal chain was analysed and experimental evidence was presented, that cathodic photodiodes have been produced. The limiting factors and the potential for application have been discussed based on supporting experiments, clearly showing the high capability of being used in a photobioenzymatic device or in a photo-switchable biosensor application.