Cavity depth and width effects on cyclophane-steroid recognition: molecular complexation of cholesterol and progesterone in aqueous solution.

BACKGROUND

Recent X-ray crystal structures show that steroid-binding proteins contain deep hydrophobic cavities defined by aromatic amino-acid side chains which encapsulate steroid molecules. These cavities resemble the binding site of a synthetic macrotricyclic cyclophane receptor which we recently reported to form complexes with cholesterol in aqueous solution. The binding affinity of the cyclophane-cholesterol complex (Ka approximately 10(6) M-1, 295 K) is similar to that measured for the cholesterol complex of steroid-transport proteins such as sterol carrier protein-2 (SCP-2). Here we describe synthesis and binding studies of a related receptor with a cavity that is wider and 2 A deeper than that of the previous cyclophane, and a comparison of the steroid-binding affinity and selectivity of the two synthetic receptors.

RESULTS

A new tricyclic cyclophane receptor with a 13 A deep cavity was synthesized to study the effect of increased cavity depth on receptor selectivity for steroids. NMR analysis demonstrated that this receptor provided increased steroidal side-chain encapsulation with a corresponding gain in binding free energy of 0.9 kcal mol-1 (in d4-methanol) as compared to our previously reported 11 A deep receptor. An unexpected consequence of the increase in cavity depth was a corresponding enlargement of the cavity width, as indicated both by steroid-binding studies and molecular modeling. This enlargement in cavity width increases binding affinity for saturated steroids while decreasing the association strength of unsaturated steroids such as cholesterol. In water, cholesterol binds to the new receptor with Ka approximately 1.5 x 10(5) M-1 and exhibits a significant complexation-mediated solubility increase.

CONCLUSIONS

Small changes in steroid receptor dimensions have resulted in large differences in steroid selectivity and binding affinity. These results indicate that potentially large gains in steroid-binding free energy may be obtainable from complete hydrophobic encapsulation of the flexible aliphatic steroidal side chain. These results have implications for the design of synthetic receptor mimics of natural steroid binding proteins.