Studies on bovine adrenal estrogen sulfotransferase. V. Synthesis and assay of analogs of 3′-phosphoadenosine 5′-phosphosulfate as cosubstrates for estrogen sulfurylation

Abstract

Analogs of adenosine 3−phosphate 5′-phosphosulfate (3′PAdo5′PS ∗ ∗ Abbreviations of the various compounds are listed in the Appendix on p. 93. ) have been prepared by an extension of a method for the synthesis of 3′PAdo5′PS, itself. Reaction of the ribonucleoside 2′,3′-cyclic phosphate 5′-phosphate with triethylamine sulfonic acid leads to a ribonucleoside 2′,3′-cyclic phosphate 5′-phosphosulfate. The latter, on treatment with ribonuclease-T 2, provides the 3′-phosphate analog. Spleen phosphodiesterase, on the other hand, promotes opening of the 2′,3′-cyclic phosphate residue to a 2′-phosphate analog. In this manner, the 2′,3′-cyclic phosphates of 8-bromoadenosine-, nebularine-, inosine-, tubercidin-, and formycin-, 5′-phosphosulfates have been converted to corresponding 2′-, and 3′-phosphate analogs. V for the utilization of 3′PAdo5′PS analogs in the sulfurylation of estrone, as mediated by bovine adrenal estrogen sulfotransferase (3′-phosphoadenylylsulfate : estrone 3-sulfotransferase, EC 2.8.2.4), is decreased more than 50% by the replacement of the 7-(ring)nitrogen atom by carbon or of the 8-H atom by bromine in the adenine moiety. However, binding of these analogs at the active site of the enzyme is still very efficient (small K m and K i values; competitive inhibition). The replacement of the 9-nitrogen atom of adenine by carbon lowers V even more, and the replacement of the 6-amino group by H, i.e., nebularine, or by 0 (inosine) results in 90 and 100% loss of enzyme activity, respectively. Shifting of the 3′-phosphate to the 2′ position in the ribose of adenosine is also detrimental and results in noncompetitive inhibition of 3′PAdo5′PS utilization.