Previously reported molecular weights for the monomeric steroid binding subunit of the androgen receptor protein have ranged from 25 000 to 167 000. The molecular weight appeared to vary among different species and target organs, as well as between different investigators. This study has examined androgen receptors from a diverse group of organs and species to determine whether these tissues share a common monomeric form. Gel filtration revealed peaks of specific [3H]dihydrotestosterone binding activity corresponding to Stokes radii of 54, 33, and 20 A in cytosols from several tissues. Phosphocellulose chromatography diminished the appearance of the smaller androgen receptor forms and facilitated the appearance of the larger 54-A form. Mixing experiments suggested that phosphocellulose was stabilizing the 54-A form by binding putative proteases which cleave this larger form. Methods were developed to generate homogeneous preparations of a given androgen receptor size for comparative study. Sucrose density gradient analysis showed sedimentation coefficients of 4.5-5.0, 3.5-4.0, and 2.5-3.0 S, respectively. The corresponding calculated molecular weights were 109 000–121 000, 52000–59000, and 22 000–27000. Scatchard analysis of each of these androgen receptor forms demonstrated very similar affinity for [3H]dihydrotestosterone (Kd ~ 1 nM), and each form possessed the ability to bind to DNA-cellulose. Extensively purified preparations of androgen receptor from R3327 tumor contained varying amounts of the three receptor forms even though molybdate and phosphocellulose were used to stabilize the androgen receptor protein during purification. It is concluded that androgen receptors from a variety of tissues share a common monomeric subunit (54 A, 4.5-5.0 S; ~ 110 000-120 000) and that stabilization is necessary during analytical and purification procedures to prevent cleavage of the monomer by endogenous proteases.
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