Defect engineering has recently emerged as a cutting-edge discipline for precise modulation of electronic structures in nanomaterials, shifting the paradigm in nanoscience from passive ‘inherent defect tolerance’ to proactive ‘defect-controlled design’. The deliberate introduction of defect—including vacancies, dopants, and interfaces—breaks the rigid symmetry of crystalline lattices, enabling new pathways for optimizing catalysis performance. This review systematically summarizes the mechanisms underlying defect-mediated electronic structure at active sites regulation, including (1) reconstruction of the electronic density of states, (2) tuning of coordination microenvironments, (3) charge transfer and localization effects, (4) spin-state and magnetic coupling modulation, and (5) dynamic defect and interface engineering. These mechanisms elucidate how defect-induced electronic restructuring governs catalytic activity and selectivity. We further assess advanced characterization techniques and computational methodologies for probing defects-induced electronic states, offering deeper mechanistic insights at atomic scales. Finally, we highlight recent breakthroughs in defect-engineered nanomaterials for catalytic applications, including hydrogen evolution reaction (HER), oxygen evolution reaction (OER) and beyond, while discussing existing challenges in scalability, defect stability, and structure–property causality. This review aims to provide actionable principles for the rational design of defects to tailor electronic structures toward next-generation energy technologies.