Enzymic isomerization of Δ⁵-3-ketosteroids

TY - JOUR

T1 - Enzymic isomerization of Δ5-3-ketosteroids

AU - Talalay, Paul

AU - Ven Shun Wang, Shun Wang

N1 - Funding Information: (a gift from Prof. L. F. FIESER TM) was not a substrate for the bacterial isomerase. The isomerization reactions were found to be complete and reversibility could not be demonstrated. The mechanism of the enzymic isomerization has been studied with the aid of D20. The enzymic reaction proceeds at identical rates in H20 and 89 % D20. Alkali catalyzed isomerization in D20 results in the incorporation of deuterium from the medium, as first shown by ANCHEL AND SCHOENHEIMER 11 for the conversion of 5-cholesten-3-one to 4-cholesten-3-one. Three samples (2.5 rag) of 5-androstene-3,I7-dione were isomerized at 25 ° respectively with 0.23 N HC1, 0.096 N NaOH and purified bacterial isomerase in a medium contammng • " 89/o o/ D20. The reactions were complete in ten minutes or less; the products were isolated, chromatographed on silica gel by gradient elution, diluted with unlabeled 4-androstene-3,I7-dione, repurified, and analyzed for deuterium in the mass spectrometer. Whereas the acid isomerized product contained 0.95 atoms D per molecule and the alkali isomerized material contained 3.86 atoms D per molecule, the enzyme isomerized product contained only o.12 atoms D per molecule. A similar experiment with TaD likewise revealed practically no incorporation of the tritium from the medium during the enzymic isomerization. In the presence of alkali the two ketonic groups are enolized and this readily accounts for the incorporation of 4 deuterium atoms TM. The enzymic mechanism suggests that there is a direct transfer of a proton from position 4 to 6 on the enzyme surface without exchange with the medium. If a direct intramolecular transfer of a proton does indeed occur, the steroid isomerase reaction provides the first recognized enzymic example. The direct and stereospecific nature of the transfer of hydrogen between substrate and coenzyme has been demonstrated in a number of enzymic reactions TM. This investigation was supported in part by a grant from the American Cancer Society on recommendation of the Committee on Growth, National Research Council. The authors wish to thank Dr. B. VENNESLAND for valuable discussion, Dr. F. LOEWUS and Mr. L. GRAVES for the deuterium analyses and Drs. GEORGE T. OKITA and HAROLD WERBIN for the tritium analyses. Copyright: Copyright 2018 Elsevier B.V., All rights reserved.

PY - 1955

Y1 - 1955

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U2 - 10.1016/0006-3002(55)90079-2

DO - 10.1016/0006-3002(55)90079-2

M3 - Article

C2 - 13276386

AN - SCOPUS:50449136029

SN - 0006-3002

VL - 18

SP - 300

EP - 301

JO - BBA - Biochimica et Biophysica Acta

JF - BBA - Biochimica et Biophysica Acta

IS - C

ER -