Isolation of dissimilar components of the 8.5S nonactivated uterine progestin receptor.
Sucrose gradient analysis of the binding of our monoclonal antibody (secreted by cell line KN 382/ECl) to [17 alpha-methyl-3H] R5020-labeled rabbit uterine progestin receptors revealed that the antibody bound only to the 8.5S form (Kd = 0.86 nM) and not to the 7S and smaller complexes. 125I-labeled antibody, on the other hand, bound to both the 8.5S complex and a component of dissociated receptor. Calculation of the relative mass (Mr) of the 125I-labeled immunoglobulin G1 (IgG1)-protein complex indicated the addition of a 60 000 Mr peptide. Electrophoretic analysis of immunoaffinity purified receptor substantiated this by revealing two protein bands (Mr approximately equal to 92 000 and Mr approximately equal to 59 000). Sequential washing of adsorbed receptor was accompanied by dissociation of the bound steroid and the Mr 92 000 peptide. The Mr approximately equal to 59 000 peptide could only be completely eluted from the immunoadsorbent under denaturing conditions. Autofluorography of receptor complexes covalently bound with [17 alpha-methyl-3H]R5020 revealed two bands, one with a Mr approximately equal to 116 000 and the second with a Mr approximately equal to 90 000. Upon immunoprecipitation both peptides precipitated with the Mr approximately equal to 92 000 and Mr approximately equal to 59 000 peptides. Gel electrophoresis demonstrated that the Mr approximately equal to 92 000 peptide and the Mr approximately equal to 90 000 did not comigrate.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D005260:Female; D007074:Immunoglobulin G / Q000276:immunology; D007163:Immunosorbent Techniques; D008970:Molecular Weight; D011487:Protein Conformation; D011817:Rabbits; D011980:Receptors, Progesterone / Q000276:immunology / Q000302:isolation & purification*; D014599:Uterus / Q000032:analysis*
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D011980:Receptors, Progesterone