Hemorrhagic enteritis virus (HEV) is an adenovirus that infects turkeys and causes immunosuppression and mortality. The virus used for the inactivated vaccine is extracted from spleens of infected turkeys, since its propagation in tissue cultures or embryonated eggs is unsuitable for mass production. The aim of this study was to develop a subunit vaccine based on a capsid protein of the virus. The knob protein, together with an adjacent part of the shaft domain pertaining to the fiber protein of HEV, was expressed in Escherichia coli and tested as a vaccine. Vaccination with this recombinant protein conferred protection against challenge in controlled and in floor-pen experiments. This finding suggests that the knob protein may be used as safe and efficient vaccine against hemorrhagic enteritis of turkeys. The possibility that the knob proteins of other adenoviruses may be protective and serve as vaccine is also discussed.