Subunit structure of rabbit testosterone estradiol-binding globulin.
Testosterone estradiol-binding globulin (rbTeBG) of rabbit plasma was purified to homogeneity by the sequential use of ammonium sulfate precipitation, affinity chromatography on 5 alpha-dihydrotestosterone-Sepharose, reactive blue-Sepharose, and finally preparative polyacrylamide gel electrophoresis. An overall purification of greater than 3000-fold was achieved with a recovery of 32%. This purified material was shown to possess the following physical characteristics: an equilibrium dissociation constant at 4 degrees C for dihydrotestosterone estimated to be 0.59 X 10(-9) M and a half-time of dissociation of rbTeBG-dihydrotestosterone complex of approximately 9 min at 4 degrees C. The purified protein shows considerable microheterogeneity with regard to size on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and to charge on isoelectric focusing gels. Analytical gel electrophoresis in the presence of SDS revealed two molecular weight components of 41,000 and 45,000. Size analysis on native gels and cross-linking studies yields a molecular weight of 101,600 and 83,000, respectively. These size components were shown to be very closely related with regard to primary structure. rbTeBG was completely absorbed by concanavalin A Sepharose resin, demonstrating the glycoprotein nature of rbTeBG. Highly purified rbTeBG and unpurified rbTeBG from rabbit plasma could be photolabeled with [3H]delta 6-testosterone and when analyzed on SDS-polyacrylamide gel electrophoresis (PAGE), two major bands of radioactivity with molecular weights of 41,000 and 45,000 appeared in both samples.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D011817:Rabbits; D012738:Sex Hormone-Binding Globulin / Q000302:isolation & purification* / Q000378:metabolism
D000596:Amino Acids; D046911:Macromolecular Substances; D012738:Sex Hormone-Binding Globulin