Amundsen Lotta K, Sirén Heli
VTT, Technical Research Centre of Finland, Espoo, Finland.
Electrophoresis. 2007 Oct;28(20):3737-44. doi: 10.1002/elps.200700271.
ACE is a popular technique for evaluating association constants between drugs and proteins. However, ACE has not previously been applied to study the association between electrically neutral biomolecules and plasma proteins. We studied the affinity between human and bovine serum albumins (HSA and BSA, respectively) and three neutral endogenous steroid hormones (testosterone, epitestosterone and androstenedione) and two synthetic analogues (methyltestosterone and fluoxymesterone) by applying the partial-filling technique in ACE (PF-ACE). From the endocrinological point of view, the distribution of endogenous steroids among plasma components is of great interest. Strong interactions with albumins suppress the biological activity of steroids. Notable differences in the association constants were observed. In the case of the endogenous steroids, the interactions between testosterone and the albumins were strongest, and those between androstenedione and the albumins were substantially weaker. The association constants, K(b), for testosterone, epitestosterone and androstenedione and HSA at 37 degrees C were 32 100 +/- 3600, 21 600 +/- 1500 and 13 300 +/- 1300 M(-1), respectively, while the corresponding values for the steroids and BSA were 18 800 +/- 1500, 14 000 +/- 400 and 7800 +/- 900 M(-1). Methyltestosterone was bound even more strongly than testosterone, while fluoxymesterone was only weakly bound by the albumins. Finally, the steroids were separated by PF-ACE with HSA and BSA used as resolving components.
亲和毛细管电泳（ACE）是一种用于评估药物与蛋白质之间结合常数的常用技术。然而，ACE此前尚未应用于研究电中性生物分子与血浆蛋白之间的结合。我们通过在亲和毛细管电泳中应用部分填充技术（PF-ACE），研究了人血清白蛋白和牛血清白蛋白（分别为HSA和BSA）与三种中性内源性甾体激素（睾酮、表睾酮和雄烯二酮）以及两种合成类似物（甲基睾酮和氟甲睾酮）之间的亲和力。从内分泌学的角度来看，内源性甾体激素在血浆成分中的分布备受关注。与白蛋白的强烈相互作用会抑制甾体激素的生物活性。观察到结合常数存在显著差异。在内源性甾体激素的情况下，睾酮与白蛋白之间的相互作用最强，而雄烯二酮与白蛋白之间的相互作用则弱得多。在37℃时，睾酮、表睾酮和雄烯二酮与HSA的结合常数K(b)分别为32100±3600、21600±1500和13300±1300 M⁻¹，而这些甾体激素与BSA的相应值分别为18800±1500、14000±400和7800±900 M⁻¹。甲基睾酮的结合甚至比睾酮更强，而氟甲睾酮仅与白蛋白有较弱的结合。最后，以HSA和BSA作为分离成分，通过PF-ACE对甾体激素进行了分离。
