STEROID RECEPTOR BINDING: A COMPARISON OF AN EMPIRICAL MODEL AND THE OBSERVED STRUCTURE

William L. Duax and Jane F. Griffin

Hauptman-Woodward Medical Research Institute, Inc. 73 High Street, Buffalo, New York 14203 USA duax@hwi.buffalo.edu

The characteristic responses of steroidal hormones require their binding to specific receptor proteins in target tissue. The existence of antagonists that compete for the steroid binding site of the receptor with high affinity demonstrates that the phenomena of binding and activity are at least partially independent. On the basis of examination of the X-ray crystal structures of compounds having high affinity for estrogen, progestin, and corticoid receptors, we proposed that receptor binding is primarily the result of a tight association between the receptor and the steroidal A ring. We postulated that agonists and antagonists that compete for a common site on a steroid hormone receptor must have A rings of identical or closely comparable composition that would account for their binding to a common site and differences in the D ring region such that features critical to induction of hormone response are present on agonists while antagonists should lack these required features andlor have other structural features that interfere with subsequent receptor functions essential to activity [1]. Our 'A-ring binding/D-ring acting' model for progestins and for estrogens, first proposed in 1978, was viewed with skepticism because the prevailing theory was that the high affinity of steroids for the receptor suggested a close lock and key fit over the entire steroid surface so that all potential hydrogen bonds and van der Waals contacts could contribute to binding.

1. The empirical model that we proposed for estrogen receptor binding and activity included the following features:
2. The C(3) hydroxyl may act as a hydrogen donor and acceptor.
3. Water may play an important role in estradiol receptor interaction.
4. A phenolic ring that can mimic the A ring of estradiol appears to be suff'icient to permit effective competition for binding to the estrogen receptor.
5. Considerable variation at the D ring region of the steroid or the corresponding region in the nonsteroidal estrogens is compatible with high affinity bind.ing and differentiation between agonist and antagonist behavior.

All of these predictions are confirmed with the recently determined crystal structures of the estrogen receptor binding domain with agonist (estradiol) and antagonist (relaxifene) bound [2].

We speculated in 1980 that the D rings of estrogens, progestins, and corticoids might control activity by:

1. Inducing or stabilizing an essential conformational state in the receptor (allostery).
2. Influencing the aggregation state of the receptor
3. Participating in a direct interaction with DNA in chromatin.

The comparison of the estrogen receptor binding domain complex revealed by X-ray structure determinations indicates that the first mechanism certainly applies. It may be too soon to eliminate proposals 2 and 3 as additional hormone functions. In an effort to fine tune the empirical model so that it could more effectively be used in drug design, we compared estrogen agonists and antagonists [3]. We considered two possible ways in which the antiestrogen LY 156758 (Relaxifene) would occupy the estrogen binding site with respect to the fit of estradiol, differing as to which of two phenolic rings in relaxifene would mimic the estradiol A ring (Figure 1).

The recent X-ray structure determination of the complexes of estradiol and LY156758 with the estrogen binding domain of the receptor indicates that the predicted it illustrated in Figure la was remarkably accurate.

Research supported by NIH Grant No. DK26546.

1. Duax, W.L., Steroid Structure and Function from X-ray Crystallographic Studies, in Molecular Structure and Biological Activity of Steroids, CRTC Press Inc., pp.1-31 (1992).
2. Brzozowski, A.M., Pike, A.C.W., Dauter, Z., Hubbard, R.E., Bonn, T., Engstrdm, O., &127hman, L., Greene, G.L., Gustafsson, J-A., and Carlquist, M., Molecular Basis of Agonism and Antagonism in the Oestrogen Receptor, Nature 389, 753 (1997).
3. Duax, W.L. and Griffin, J.F., Steroid Molecular Structure, Receptor Binding and Hormone Action, in Proceedings of 2nd Int'l. Center for Biotechnology Symposium: The Steroid/Thyroid Hormone Receptor Family and Gene Regulator, J. Carlated-Duke, H. Eriksson &. J.A. Gustafsson (Eds.), Birkhauser Verlag, Basel, Boston, Berlin, pp. 319-335 (1989).