A novel synthetic approach for the electrospinning of CuO nanofibers was applied to produce fibrous CuO photocathodes, calcined at different temperatures, in order to evaluate the impact of crystallinity and defect sites on solar-driven pho-toelectrochemistry. By careful optimization of the preparation conditions, stable electrospinning was achieved, allowing the fabrication of large quantities of highly crystalline, uniform CuO nanofibers. The as-spun fibers were calcined at 300 degrees C, 400 degrees C, 550 degrees C, and 800 degrees C in air, and their crystallographic and structural evolution was systematically characterized. The correlation of the physicochemical properties with the photoelec-trochemical performance of CuO nanofiber photocathodes re-veals a structure-property relationship: The higher the annealing temperature, the more developed are the crystalline domains of the nanofibers, which in turn result in better conductivity and less defect sites serving as trap states for the photoexcited charge carriers. Thus, the CuO nanofiber photocathodes annealed at 800 degrees C showed the highest photoresponse and stability of the photocathodes reported herein, with no loss of photocurrent after prolonged operation in an aqueous electrolyte. Further improvement of the CuO photocathodes was realized by increasing the film thickness leading to photocurrents up to 0.16 mA at 0.4 V vs. RHE, however the stability of the thick photocathode remains a critical issue.