Semi-artificial photosynthesis integrates photosynthetic enzymes with artificial electronics, which is an emerging approach to reroute the natural photoelectrogenetic pathways for sustainable fuel and chemical synthesis. However, the reduced catalytic activity of enzymes in bioelectrodes limits the overall performance and further applications in fuel production. Here, we show new insights into factors that govern the photoelectrogenesis in a model system consisting of photosystem II and three-dimensional indium tin oxide and graphene electrodes. Fluorescence microscopy and in situ surface-sensitive infrared spectroscopy are employed to probe the enzyme distribution and penetration within electrode scaffolds of different structures, which is further correlated with protein film-photoelectrochemistry to establish relationships between the electrode structure and enzyme activity. We find that the hierarchical 1 structure of electrodes mainly affects the protein integration, but not the enzyme activity. Photoactivity is more limited by light intensity and electronic communication at the biointerface. This study provides guidelines for maximizing the performance of semi-artificial photosynthesis and also presents a set of methodologies to probe the photoactive biofilms in three-dimensional electrodes.