Abstract
The binding of estradiol-17 beta (E2), diethylstilbestrol (DES), and polyene fatty acids, in particular arachidonate (C20:4), to alpha 1-fetoprotein (alpha-FP) and albumin purified from mouse embryo sera was studied using equilibrium dialysis and electrophoretic techniques. E2, arachidonate, and DES all bind to alpha-FP, but with decreasing strength. E2 is a high affinity, low capacity ligand (Ka approximately 0.8 X 10(8) M-1 and approximately 0.3 sites/mol of alpha-FP at 25 degrees C); arachidonate is a weaker ligand disposing of more sites (Ka approximately 0.3 X 10(7) M-1 and 4-5 sites/mol of alpha-FP); the binding of DES is of comparatively low affinity and capacity (Ka approximately 0.2 X 10(7) M-1 and n approximately 0.7/mol of alpha-FP). In spite of different structures and equilibrium parameters, E2, DES, and arachidonate are able to compete with each other for binding to the fetoprotein. The C22:4 and C22:6 fatty acids are also efficient concentration-dependent inhibitors of E2 or DES binding. Albumin binds the fatty acids and DES, but equilibrium parameters are different from those of alpha-FP. In particular, arachidonate is a better ligand for albumin, where it interacts with at least two classes of apparent sites (Ka1 approximately 0.3 X 10(8) M-1 and n1 approximately 1; Ka2 approximately 0.2 X 10(7) M-1 and n2 approximately 30). In contrast to alpha-FP, albumin virtually does not bind E2. Also, no competition could be demonstrated between DES and fatty acid ligands for binding to albumin. None of the studied interactions, with either albumin or alpha-FP, was modified even by high doses of bilirubin. The possible functions of the various binding activities present in fetal sera in the process of growth are discussed.
Highlights
Ate (C20:4), to al-fetoprotein (a-FP) and albumin puri- Along with the rat, the mouse is the only species known to fied from mouse embryo serawas studied using equi- date which displays higher steroid-protein interactions in the librium dialysis and electrophoretic techniques
In We report on the application of equilibrium dialysis and particular, arachidonateis a better ligand for albumine,lectrophoretic techniques to measure the interactions of natwhere it interacts withat least two classes of apparent ural and syntheticligands-in particular estradiol-l7P, dieth
We measured the labeling of the and the inertness of its albumin towards the estrogens, with two proteins after polyacrylamide gel electrophoresis of fetal theother testedinteractionsshared by the two proteins, sera, preincubated with the different tritiated ligands
Summary
Ate (C20:4), to al-fetoprotein (a-FP) and albumin puri- Along with the rat, the mouse is the only species known to fied from mouse embryo serawas studied using equi- date which displays higher steroid-protein interactions in the librium dialysis and electrophoretic techniques. Interactions of a-FPwith Arachidonic Acidand Diethylstilbestrol: Comparison to the Binding of Estradiol-17p-C values, association constants (ICa),and number of binding sites (n) were measured at equilibrium for the interactions of purified a-FP with the natural estrogen, the polyene fatty acid, and the estrogen analogue.
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