In this study, lipoxygenase (LOX) extracted from dry-cured mackerel was purified, resulting in a 4.1-fold purification factor with a specific activity of 493.60 U/min·g. LOX enzymatic properties were assessed, referring to its optimal storage time (1–2 days), temperature (30 °C), and pH value (7.0). The autoxidation and LOX-induced oxidation of palmitic acid (C16:0), stearic acid (C18:0), oleic acid (C18:2n9c), linoleic acid (C18:2n6c), arachidonic acid (C20:4), EPA (C20:5), and DHA (C22:6n3) were simulated to explore the main metabolic pathways of key flavors in dry-cured mackerel. The results showed that the highest LOX activity was observed when arachidonic acid was used as a substrate. Aldehydes obtained from LOX-treated C18:1n9c and C18:2n6c oxidation, which are important precursors of flavors, were the most abundant. The key flavors in dry-cured mackerel were found in the oxidative products of C16:0, C18:0, C18:1n9c, C18:2n6c, and C20:4. Heptanaldehyde could be produced from autoxidation or LOX-induced oxidation of C18:0 and C18:1n9c, while nonal could be produced from C18:1n9c and C18:2n6c oxidation. Metabolic pathway analysis revealed that C18:1n9c, C18:2n6c, EPA, and DHA made great contributions to the overall flavor of dry-cured mackerel. This study may provide a relevant theoretical basis for the scientific control of the overall taste and flavor of dry-cured mackerel and further standardize its production.