It remains a vital task to establish ultrasensitive sensing interfaces for detection of target analytes to meet the demands of modern analysis. Herein, a highly sensitive turn-on photoelectrochemical (PEC) platform for trace 17β-estradiol (E2) assay was developed based on Au nanrods (AuNRs) with surface plasmon resonance (SPR) properties induced signal amplification. Specifically, a ternary hybrid was prepared by integrating hematite (α-FeO) nanocrystals and N-doped graphene (NG) with AuNRs, which further served as highly efficient photoactive species. Subsequently, a PEC sensing platform was fabricated based on the specific binding of E2 and its aptamer. On such a sensor, the capture of E2 molecules by aptamers led to increased photocurrent. This was attributed to that the specific recognition reaction between E2 and aptamer resulted in the conformational change of the aptamers and complete dissociation of some aptamers on the PEC sensing interface. It can be confirmed by the electrochemical impedance spectroscopy (EIS) results. This process decreased the steric hindrances between the electrode surface and solution and thus increased the photocurrent response. Under the optimal conditions, the as-prepared PEC aptasensor exhibited superb analytical performances for detection of E2 in the range from 1×10M to 1×10M with a detection limit of 3.3×10M. The aptasensor manifested outstanding selectivity towards E2 when other endocrine disrupting compounds with similar structure coexisted. Furthermore, the aptasensor was successfully applied for the determination of E2 in milk powder. The present strategy provides a potential way to boost the activity of photoactive materials and improve the sensitivity of PEC biosensor.