Characterization and measurement of the plasma alpha- and beta-sex hormone-binding globulin paralogs in salmon.

When the biochemical characteristics of coho salmon SHBG (csSHBG) plasma were examined, two different steroid-binding profiles were obtained corresponding to recombinant csSHBG-alpha and csSHBG-beta. These SHBG paralogs share only 24% sequence identity, and this explains their unique steroid-binding properties. Both proteins bind testosterone, but csSHBG-alpha also binds androstenedione (Kd = 2.8 nm) and ethinylestradiol with high affinity, whereas csSHBG-beta binds estradiol (Kd = 0.8 nm) preferentially. When analyzed by gel filtration, csSHBG-alpha displays the properties of a 153-kDa homodimer, whereas csSHBG-beta elutes as a 68-kDa monomer. The unique steroid-binding properties of csSHBG-alpha and csSHBG-beta allowed us to develop an assay for their measurements in immature (pre-smolt) and mature coho salmon blood. Plasma csSHBG-alpha levels were 3- to 4-fold higher than those of csSHBG-beta irrespective of developmental stage or sex and correlate with each other. The major site of csSHBG-alpha expression in pre-smolts and mature fish is the liver, but low levels of csSHBG-alpha mRNA are present in stomach/intestine of mature fish. In pre-smolts, high levels of csSHBG-beta mRNA are present in gills and ovary, whereas csSHBG-beta mRNA is most abundant in muscle and stomach/intestine of mature fish. Based on the differences in csSHBG-alpha and csSHBG-beta plasma levels and their tissue expression profiles, we conclude that gills and/or muscle contribute mainly to plasma SHBG-beta in coho salmon. The assays we have developed will enable studies of how SHBG-alpha/SHBG-beta biosynthesis is regulated throughout the salmonid life cycle and how they influence steroid hormone action in these fish.