Nattokinase is a thrombolytic enzyme obtained from Japanese traditional food natto for prevention and cure of thrombosis-related cardiovascular diseases. However, the effectiveness of nattokinase through oral intake is limited, due to the loss of thrombolytic activity in the acidic gastric juice. In this study, we develop a functional oral delivery system of nattokinase, in which chitosan microparticles were used as the carrier core to load nattokinase via genipin crosslinking and then covered by a casein-based protective shell via transglutaminase (TG) crosslinking. The results of in vitro and in vivo assays, in the aspects of bioactivity, release dynamics, and therapeutic effects, indicated that the bilayer shell-core structure could protect loaded nattokinase from destruction in the gastric juice and achieve its controlled-release in the intestine. This work demonstrates the availability of bilayer shell-core structure design in oral delivery of nattokinase and shows its high potential for application as an anti-thrombosis functional food additive.