TY - JOUR
T1 - Immunochemical analysis of the glucocorticoid receptor
T2 - Identification of a third domain separate from the steroid-binding and DNA-binding domains
AU - Carlstedt-Duke, J.
AU - Okret, S.
AU - Wrange, O.
AU - Gustafsson, J. A.
PY - 1982
Y1 - 1982
N2 - The glucocorticoid-receptor complex can be subdivided into three separate domains by limited proteolysis with trypsin or α-chymotrypsin. The following characteristics can be separated: steroid-binding activity (domain A), DNA-binding activity (domain B), and immunoactivity (domain C). We have previously reported the separation of the steroid-binding domain from the DNA-binding domain by limited proteolysis of the receptor with trypsin. In this paper, we report the detection by immunochemical analysis of a third domain of the glucocorticoid receptor, which does not bind hormone. Immunoactivity was detected by using specific antiglucocorticoid receptor antibodies raised in rabbits against purified rat liver glucocorticoid receptor and the assay used was an enzyme-linked immunosorbent assay. After digestion with α-chymotrypsin, the immunoactive region of the receptor (domain C) was separated from the other two domains (A and B). The immunoactive fragment was found to have a Stokes radius of 2.6 nm. Further digestion with α-chymotrypsin resulted in separation of the immunoactive fragment to give a fragment having a Stokes radius of 1.4 nm. The immunoactive domain could be separated from the half of the glucocorticoid receptor containing the steroid-binding and the DNA-binding domains (Stokes radius, 3.3 nm), by limited proteolysis of the receptor by α-chymotrypsin followed by gel infiltration or chromatography on DNA-cellulose.
AB - The glucocorticoid-receptor complex can be subdivided into three separate domains by limited proteolysis with trypsin or α-chymotrypsin. The following characteristics can be separated: steroid-binding activity (domain A), DNA-binding activity (domain B), and immunoactivity (domain C). We have previously reported the separation of the steroid-binding domain from the DNA-binding domain by limited proteolysis of the receptor with trypsin. In this paper, we report the detection by immunochemical analysis of a third domain of the glucocorticoid receptor, which does not bind hormone. Immunoactivity was detected by using specific antiglucocorticoid receptor antibodies raised in rabbits against purified rat liver glucocorticoid receptor and the assay used was an enzyme-linked immunosorbent assay. After digestion with α-chymotrypsin, the immunoactive region of the receptor (domain C) was separated from the other two domains (A and B). The immunoactive fragment was found to have a Stokes radius of 2.6 nm. Further digestion with α-chymotrypsin resulted in separation of the immunoactive fragment to give a fragment having a Stokes radius of 1.4 nm. The immunoactive domain could be separated from the half of the glucocorticoid receptor containing the steroid-binding and the DNA-binding domains (Stokes radius, 3.3 nm), by limited proteolysis of the receptor by α-chymotrypsin followed by gel infiltration or chromatography on DNA-cellulose.
UR - http://www.scopus.com/inward/record.url?scp=0020328733&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0020328733&partnerID=8YFLogxK
U2 - 10.1073/pnas.79.14.4260
DO - 10.1073/pnas.79.14.4260
M3 - Article
C2 - 6181503
AN - SCOPUS:0020328733
VL - 79
SP - 4260
EP - 4264
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
SN - 0027-8424
IS - 14 I
ER -