Potent Inhibition Of Human SULT1A1 By 17α-Ethinylestradiol

Potent Inhibition Of Human SULT1A1 By 17α-Ethinylestradiol (EE2). Sulfotransferases (SULTs) have a major role in metabolizing endogenous and exogenous compounds in the human body. SULTs are Phase II conjugating enzymes that activate or inactivate compounds by transferring a sulfonate group from 3’-phosphoadenosine 5’-phosphosulafate (PAPS) to the hydroxyl or amine group. Human SULT1A1 is highly expressed in liver and is also widely expressed in other tissues including brain, breast and intestine where it is associated with the sulfation of a wide variety of small phenols, dietary polyphenols, thyroid hormones and N-hydroxyaromatic amines. SULT1A1 also conjugates estrogens including 17α-ethinylestradiol (EE2), the active ingredient in many oral contraceptives. EE2 is a lipophilic molecule that passes the outer membrane of the cell and targets estrogen receptors to mediate hormone-regulated gene expression. SULT1A1 sulfates EE2 with a Km of approximately 600 nM. This study focused on the ability of EE2 to potently inhibit the SULT1A1 catalyzed sulfation of 17β-estradiol (E2), p-nitrophenol, and β-naphthol. The Ki values ranged from 8.9 to 19 nM for the inhibition of the SULT1A1 catalyzed sulfation of these compounds. The Ki for EE2 inhibition of E2 sulfation by intact human MCF-7 breast cancer cells that express high levels of SULT1A1 activity was 20 nM. We were interested in the mechanism by which EE2 could inhibit SULT1A1 at low nanomolar concentrations while being a substrate at greater than 30-fold higher concentrations. PAP(S) binding results in a structural rearrangement that decreases the volume of the PAPS and substrate binding sites. Models of SULT1A1 were generated with the Molecular Operating Environment programs using the crystal structures of human SULT2A1 resolved with bound PAPS (closed) and of the enzyme without bound PAPS (open) as a template. The Kd for EE2 binding to pure SULT1A1 was determined to be 150 nM to the enzyme without bound PAP and 4.3 nM to the enzyme previously incubated with PAP. EE2 docked into the active site of the SULT1A1 open model in a catalytically competent orientation with low affinity, whereas, if PAPS was bound to SULT1A1 to generate a closed conformation, EE2 bound with a greater affinity in a non-catalytically competent orientation. This demonstrates that when EE2 is bound to SULT1A1 before PAPS is bound, EE2 is in a catalytically orientation for sulfation; however, when PAPS is bound to SULT1A1 first EE2 acts as a non-sulfated potent competitive inhibitor. The potent inhibition of SULT1A1 by low nanomolar EE2 concentrations may be a source of drug interactions in women using EE2 containing contraceptives. (Supported by NIH GM38953)