The effects of various serine protease inhibitors on estrogen receptor steroid binding.
Two serine protease inhibitors, phenylmethanesulfonyl fluoride (PMSF) and diisopropylfluorophosphate (DFP), were utilized to investigate the possible involvement of serine hydroxyl groups on 17 beta-estradiol binding to the rat estrogen receptor (ER). Single point saturation analysis and Scatchard analysis demonstrated that both 5 mM PMSF and 5 mM DFP were able to inhibit steroid binding to the ER after incubation at 37 degrees C, but neither were able to inhibit steroid binding of the nonactivated ER (0-4 degrees C). The reducing agent dithiothreitol (DTT) was used to differentiate between the interaction of PMSF with serine groups or with sulfhydryl groups of the receptor. When incubated in the presence of 5 mM PMSF, various concentrations of DTT up to 25 mM were not able to overcome the inhibition of this agent, indicating that there was no interaction of PMSF with sulfhydryl groups. Thus, these findings indicate that serine hydroxyl groups are involved in steroid binding of the rat ER.
Koffman
B
B
Department of Biochemistry, Baltimore College of Dental Surgery, University of Maryland 21201.
Modarress
K J
KJ
Bashirelahi
N
N
D000818:Animals; D003600:Cytosol / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D004958:Estradiol / Q000378:metabolism; D005260:Female; D066298:In Vitro Techniques; D007531:Isoflurophate / Q000494:pharmacology; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D011960:Receptors, Estrogen / Q000187:drug effects* / Q000378:metabolism; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology*
D011960:Receptors, Estrogen; D015842:Serine Proteinase Inhibitors; D007531:Isoflurophate; D004958:Estradiol; D010664:Phenylmethylsulfonyl Fluoride; D004229:Dithiothreitol