Phyllis D. Strong

Steroid structure and function VII. Remarkable estrogenicity of 3-hydroxy-9β-estra-1,3,5(10)-triene-11,17-dione

Remarkably high estrogenic activity was observed for 3-hydroxy-9 beta-estra-1,3,5(10)-triene-11,17-dione despite its unusual bent conformation. The 9 alpha epimer of this compound has markedly less activity despite the fact that its overall shape is nearly identical to that of estrone. The potency of these compounds in enhancing uterine weight in Fischer rats and reducing ovarian weight in parabiosed rats was compared with that of estrone, and the structures were unambiguously identified by X-ray crystallographic study. The results underscore the importance of the phenolic ring A to estrogenic activity, and suggest a tolerance of the putative estrogenic receptor to flexibility in overall molecular shape.

Steroid structure and function. Molecular conformation of 4-hydroxyestradiol and its relation to other catechol estrogens

Hydroxylation of estrogens at C(2) or C(4) effects differentially their binding affinity to and dissociation rate from the estrogen receptor. The X-ray crystal structure of 4-hydroxyestradiol (4-OH-E2) is reported here and compared with that of 2-hydroxyestradiol (2-OH-E2), the 2- and 4-hydroxylated derivatives of estrone (E1) and with that of the parent estrogens, E1 and E2. The overall molecular shape and hydrogen bonding patterns of each were examined for their possible relevance to their binding to the estrogen receptor and their biological activity. A shift in the B-ring conformation away from the symmetrical 7 alpha,8 beta-half-chair form toward the 8 beta-sofa form is induced by both 2- and 4-hydroxy substitution. This shift appears to be larger in the case of E2 than E1 derivatives and to be correlated with an observed change in the hydrogen bonding potential of the C(3) hydroxyl. In 4-OH-E2, as in E2 and 4-OH-E1, the C(3) hydroxyl functions both as a hydrogen bond donor and acceptor. In contrast in 2-OH-E2 the hydroxyl functions only as a donor. The markedly reduced affinity of 2-hydroxylated estrogens for the estrogen receptor could be due to a combination of steric interactions, competition between O(2) and O(3) for hydrogen bonds for a common site on the receptor, and to general interference with hydrogen bond formation of O(3). The C(4) hydroxyl participates in the formation of a chain of hydrogen bonds in the solid state that is similar to a chain seen in single crystals of E2. The presence of a similar chain of hydrogen bonds involving O(3) in the receptor site could account for the decreased dissociation rate of the 4-OH-E2 receptor complex.

Steroid structure and function—IX. Molecular conformation and receptor binding of isomeric analogs of d-Homo-Estradiol

Abstract The fact that rac - d -homo-8α-estra-1,3,5(10)-triene-3,17a-diol (DH8α) has a higher affinity for the estrogen receptor in rabbit uterus than rac - d -homo-estra-1,3,5(10)-triene-3,17a-diol (DH8β) may be due to differences in hydrogen bonding potential of the C(3) hydroxyl due to conformational transmission effects. The crystal structure determinations of DH8α and DH8β reported here unambiguously establish the relative configurations of the two structures and show that the difference in the binding affinity is not due to a change in O(3)–O(17) distance. The greater affinity of DH8α, estradiol, and diethylstilbestrol (DES) may be linked to an enhanced potential to act as hydrogen bond acceptors as well as donors in the active site. Crystallographic evidence suggests that the weaker binding DH8β and estrone act as donors only. Quantum calculations suggest that the energy of a sequential pair of hydrogen bonds is not merely additive (≅6 Kcal/mol) but may be as much as 9 Kcal/mol. These results support the hypothesis that there is a highly specific association between the A-rings and the binding site of the estrogen receptor protein involving O(3) as a hydrogen bond donor and acceptor and that the receptor accommodates greater variability in the d -ring region.

Steroid structure and function V. A-ring conformation in 17-hydroxy-6α-methylprogesterone

Abstract The molecular conformation of 17-hydroxy-6α-methylprogesterone has been determined crystallographically and is compared with 17-hydroxy-progesterone, 17-acetoxyprogesterone and 17-acetoxy-6α-methylprogesterone (MPA). The analysis demonstrates that the 6α-methyl substituent is not sufficient by itself to induce inversion of the A-ring. Consequently, the inverted form observed in MPA and proposed to be responsible for high affinity binding to the progesterone receptor appears to be induced by the combined long range influence of 17α-acetoxy substituent and the direct interaction of the 6α-methyl group with the flexible A-ring.

The molecular structure of 16α-iodo-17β-estradiol, a high affinity ligand for the estrogen receptor

Abstract 16α-Iodoestradiol is an estrogenic steroid with high affinity for the estrogen receptor. When labelled with a gamma emitting isotope, such as 125 I, the resulting radioactive steroid is an excellent ligand for the sensitive analysis of the estrogen receptor. The X-ray study of the crystal structure of 16α-iodoestradiol reveals that the bond distances and angles of the iodinated estrogen are similar to those of estradiol and estriol. The substitution of iodine for the 16α-hydroxyl group has little effect on geometric and electronic properties of the molecule and does not interfere with the hydrogen bonding ability of the 17β-hydroxyl group. The difference in receptor binding affinity between 16α-estradiol and estriol may be due to competition for intermolecular hydrogen bond formation between the hydroxyls at C(16) and C(17) on estriol. X-Ray data indicate that the 16-hydroxyl can form significantly shorter and presumably stronger hydrogen bonds.

Synthesis and characterization of the anomeric pair of 17β-glucuronides of ethynylestradiol

Abstract The α- and β-anomers of the 17β- d -glucuronide conjugate of ethynylestradiol were synthesized by the SnCl4-promoted reaction between β-acetoxy GAM and the t-17β-hydroxyl group of EE2-3-acetate. The conjugates were resolved by crystallization and HPLC. Positive identification was established by u.v. spectrophotometry, i.r. and mass spectrometry and 1H- and 134C-n.m.r. The structure of the β-anomer was confirmed by X-ray crystallographic analysis. In addition, the α-anomer was refractory to hydrolysis by bovine β-glucuronidase, establishing a biochemical difference between the conjugate pair.

Steroid structure and function—IX. molecular conformation of catechol estrogens

Abstract The catechol estrogens have varying degrees of affinity for the estrogen receptor, catecholamine enzymes and the dopamine receptor. The X-ray crystal structures of 2-hydroxyestradiol, 2-hydroxyestrone and 4-hydroxyestrone are reported here and compared with those of estradiol, estrone, dopamine and apomorphine. The overall molecular shapes and hydrogen bonding patterns have been examined for possible relevance to protein binding and activity. The principal structural observations and their potential significance are the following: (1) The 4-hydroxyestrone results indicate that the 4-substituent does not constitute a steric impediment to estrogen receptor binding but does restrict the probable location of the hydrogen bond acceptor atom on the receptor. (2) The reduced affinity of 2-hydroxyestradiol for the estrogen receptor may be due to a combination of steric interaction and interference with hydrogen bond formation of O(3). (3) The reduced affinity of 17-one substituted steroids in this series for the estrogen receptor most strongly supports the probable importance of a hydrogen bond donor at that position interacting directly with the receptor. (4) In the catechol estrogens the formation of an intramolecular and two or more intermolecular hydrogen bonds together with the extension of the A rings planar surface to include the hydroxyl substituents and part of the B ring must account for its high binding affinity for catechol amine related enzymes, the dopamine receptor and a putative catechol estrogen receptor. (5) Two orientations of the catechol estrogens relative to that of dopamine when bound to its receptor are suggested by this study. One achieves maximum overlap of the hydrogen bond pattern of the dihydroxy benzene ring and the other maximizes similarity in overall shape. The results of this investigation are consistent with the hypothesis that the A-ring plays a primary role in initiating estrogen binding to its receptor and suggests an even more highly specific A-ring interaction with the dopamine receptor and catechol amine enzymes.

Crystal structures of 4,11-pregnadiene-3,20-dione and 21-methyl-20-oxa-4-pregnene-3,20-dione

The structures of the title compounds were solved by direct methods and refined by anisotropic full-matrix least-squares methods. 4,11-Pregnadiene-3,20-dione, C21H28O2(1) crystallizes in the monoclinic space groupP21 (Z=2). The unit cell parametersa, b, c (Å), and β (°) were: 12.319(2), 7.700(2), 9.717(2), 109.41(2). TheA- andC-rings exhibit intermediate sofa-halfchair conformations. TheB-ring has a chair conformation and theD-ring assumes an intermediate envelope-half-chair conformation. The progesterone side chain has a typical conformation; the C16−C17−C20−O20 torsion angle is −15.1(4)°. 21-Methyl-20-oxa-4-pregnene-3,20-dione, C21H30O3 (2) crystallizes in the orthorhombic space groupP212121 (Z=4). The unit cell parametersa, b, c (Å) were: 12.926(2), 19.447(4), 7.313(1). The progesterone side chain has an unusual conformation; the C16−C17−C20−O20 torsion angle is 174.6(4)°. TheA-ring has a 1α, 2β-half-chair conformation, ringsB andC exhibit chair conformations and ringD is in a 13β,14α-half-chair conformation.

Structure of 9β-estrone

: 3-Hydroxy-9 beta-estra-1,3,5(10)-trien-17-one, C18H22O2, Mr = 270.4, monoclinic, P2(1), a = 9.527 (2), b = 11.182 (3), c = 7.078 (1) A, beta = 108.45 (1) degrees, V = 715.3 (3) A3, Z = 2, Dx = 1.255 g cm-3, lambda (Mo K alpha) = 0.71073 A, mu = 0.746 cm-1, F(000) = 292, T = 298 K, R = 0.037 for 2128 reflections with F greater than 2.0 sigma (F). The compound was one in a series of 9 beta-estrone analogues synthesized to study their estrogenic activity. The B ring conformation is a 7 beta,8 alpha-half chair, due to the configuration at C9. The O3 hydroxy forms a hydrogen bond to O17 at a distance of 2.76 A.

Structure of 3α,12β-dihydroxy-2β-morpholino-5α-pregnan-20-one, C25H41O4N

AbstractThe crystal and molecular structure of 3α,12β-dihydroxy-2β-morpholino-5α-pregnan-20-one, C25H41O4N, has been determined:Mr=419.6,P21,a=13.5778(8),b=14.4340(8),c=5.8943(5) Å,β=94.32(1)°,Vc=1151.9(3) Å3,Z=2,Dx=1.21 g cm−3,