A Further Study of a Testosterone-binding Component of Human Pregnancy Serum

Equilibrium constants of association were determined by a technique based on the principle of equilibrium dialysis but utilizing Sephadex G-25 for the strong binding exhibited by human pregnancy serum (or by the material precipitated from late pregnancy serum by ammonium sulfate at 50% saturation) for: (a) testosterone-1,2-3H, (b) radioinert 17β-estradiol (in competition with testosterone-1,2-3H), and (c) 17β-estradiol-6,7-3H. The respective values for the intrinsic association constant, k, were all quite high: (a) 4.5 × 108M−1, (b) 1.35 × 108M−1, and (c) 0.4 × 108M−1. These findings are in accord with the view that the testosterone-binding component of pregnancy serum also binds 17β-estradiol but not as strongly. The binding of 17β-estradiol by bovine serum albumin, measured by the same technique, gave nk = 9 × 104M−1. Reagents which react with —SH groups, and other reagents which reduce the disulfide bonds, depressed in striking fashion the testosterone-binding activity of pregnancy serum. It is suggested that the testosterone-binding component contains both cysteine and cystine, and that their maintenance intact is important for steroid-binding activity.