The engineering of renewable and sustainable protein-based light-to-energy converting systems is an emerging field of research. Here, we report on the development of supramolecular light-harvesting electrodes, consisting of the redox protein cytochrome c working as a molecular scaffold as well as a conductive wiring network and photosystem I as a photo-functional matrix element. Both proteins form complexes in solution, which in turn can be adsorbed on thiol-modified gold electrodes through a self-assembly mechanism. To overcome the limited stability of self-grown assemblies, DNA, a natural polyelectrolyte, is used as a further building block for the construction of a photo-active 3D architecture. DNA acts as a structural matrix element holding larger protein amounts and thus remarkably improving the maximum photocurrent and electrode stability. On investigating the photophysical properties, this system demonstrates that effective electron pathways have been created.

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The binding of photosystem I (PS I) from to the native cytochrome (cyt) and cyt from horse heart (cyt) was analyzed by oxygen consumption measurements, isothermal titration calorimetry (ITC), and rigid body docking combined with electrostatic computations of binding energies. Although PS I has a higher affinity for cyt than for cyt, the influence of ionic strength and pH on binding is different in the two cases. ITC and theoretical computations revealed the existence of unspecific binding sites for cyt besides one specific binding site close to P Binding to PS I was found to be the same for reduced and oxidized cyt Based on this information, suitable conditions for cocrystallization of cyt with PS I were found, resulting in crystals with a PS I:cyt ratio of 1:1. A crystal structure at 3.4-Å resolution was obtained, but cyt cannot be identified in the electron density map because of unspecific binding sites and/or high flexibility at the specific binding site. Modeling the binding of cyt to PS I revealed a specific binding site where the distance and orientation of cyt relative to P are comparable with cyt from purple bacteria relative to P This work provides new insights into the binding modes of different cytochromes to PS I, thus facilitating steps toward solving the PS I-cyt costructure and a more detailed understanding of natural electron transport processes.

A precursor-based approach has been employed for the construction of scalable, transparent 3D photobioelectrodes based on PSI and cyt c. An improved transparancy and high photocurrents can be achieved as compared to nanoparticle-based preparation methods.

Trimeric photosystem I from the cyanobacterium Thermosynechococcus elongatus ( Te PSI) is an intrinsic membrane protein, which converts solar energy into electrical energy by oxidizing the soluble redox mediator cytochrome c 6 (Cyt c 6 ) and reducing ferredoxin. Here, we use cryo-electron microscopy and small angle neutron scattering (SANS) to characterize the transient binding of Cyt c 6 to Te PSI. The structure of Te PSI cross-linked to Cyt c 6 was solved at a resolution of 2.9 Å and shows additional cofactors as well as side chain density for 84% of the peptide chain of subunit PsaK, revealing a hydrophobic, membrane intrinsic loop that enables binding of associated proteins. Due to the poor binding specificity, Cyt c 6 could not be localized with certainty in our cryo-EM analysis. SANS measurements confirm that Cyt c 6 does not bind to Te PSI at protein concentrations comparable to those for cross-linking. However, SANS data indicate a complex formation between Te PSI and the non-native mitochondrial cytochrome from horse heart (Cyt c HH ). Our study pinpoints the difficulty of identifying very small binding partners (less than 5% of the overall size) in EM structures when binding affinities are poor. We relate our results to well resolved co-structures with known binding affinities and recommend confirmatory methods for complexes with K M values higher than 20 μM.