Interaction of glucocorticoid receptor from rat liver with protamine and arginine.
The nontransformed glucocorticoid receptor (GR) from rat liver was found to bind to protamine-Sepharose and could be recovered by a salt gradient without a change in molecular configuration. The nontransformed GR also bound to arginine-Sepharose, but the transformed GR did not bind to either resin. Ligand-free GR interacted with both resins and was eluted without loss of its steroid binding ability. The bindings of GR to protamine- and arginine-Sepharose were saturable. The apparent dissociation constants of GR on protamine-Sepharose varied from 0.34 nM (-molybdate) to 0.68 nM (+ 10 mM molybdate) and those on arginine-Sepharose were 1.99 nM (-molybdate) and 0.65 nM (+ 10mM molybdate), respectively. The maximum binding capacity was achieved by arginine-Sepharose in the absence of molybdate. Higher salt concentrations (0.5 M NaCl) were required to elute GR from protamine-Sepharose than from arginine-Sepharose (approx 0.03 M NaCl). However, the effectiveness of several salts for the elution of GR was consistent in both resins as follows; MgCl2 = CaCl2 = Na2WO4 greater than (NH4)2SO4 = Na2MoO4 greater than arginine-HCl greater than lysine-HCl greater than KCI = NaCl. These results suggest that GR interacts with arginine residues in protamine. Chromatography using these resins resulted in 7-10-fold purification of occupied and unoccupied nontransformed GRs.
Nemoto
T
T
Department of Biochemistry, Iwate Medical University School of Dentistry, Japan.
Ohara-Nemoto
Y
Y
Ota
M
M
D000818:Animals; D001120:Arginine / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004347:Drug Interactions; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D008982:Molybdenum / Q000494:pharmacology; D011479:Protamines / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D011965:Receptors, Glucocorticoid / Q000032:analysis* / Q000187:drug effects; D012685:Sepharose / Q000031:analogs & derivatives
D011479:Protamines; D011965:Receptors, Glucocorticoid; C035771:Sepharose CL 4B; D008982:Molybdenum; D012685:Sepharose; C024687:sodium molybdate(VI); D001120:Arginine