Androgen receptor mRNA was translated in vitro, and androgen- and antiandrogen-bound receptor complexes were studied using limited proteolytic digestion by trypsin. Partial proteolysis of androgen-bound receptor protein resulted in a 29-kDa proteolysis-resisting fragment, whereas antiandrogen binding stabilised a 35-kDa fragment. Both fragments contain the entire ligand binding domain, and the 35-kDa fragment extended into the hinge region of the receptor. Several antiandrogens show agonistic properties for a mutated androgen receptor (LNCaP cell variant); trypsin digestion of antiandrogen-bound mutated receptor also resulted in a 29-kDa fragment. Our results point to an important difference between antiandrogens and antagonists of other steroid hormone receptors. Antiandrogens result in protection of both the hinge region and C-terminus of the androgen receptor agonist proteolytic attack, whereas other studies showed that antiestrogens and antiprogestagens expose the C-terminal end of the ligand binding domain of their respective receptors to protease. Differences in conformation of the hinge region distinguish androgen-bound from antiandrogen-bound receptor complexes, which represents an important feature of antiandrogen action.