INHIBITION OF GAMMA-BUTYROBETAINE HYDROXYLASE BY CYCLOPROPYL-SUBSTITUTED GAMMA-BUTYROBETAINES

Butyrobetaine hydroxylase (4-(trimethylamino)butyrate, 2-oxoglutarate: oxygen oxidoreductase (3-hydroxylating); EC 1.14.11.1) catalyzes the final step in the biosynthesis of carnitine. A stepwise, homolytic mechanism has been proposed (Englard, S.; et al. Biochemistry 1985, 24, 1110). Cyclopropyl-substituted substrate analogues 1, 2, and 3 were synthesized and evaluated as inhibitors of the enzyme with a view toward their potential as “free radical clock” probes for radical transients. Betaines 1 and 3 are competitive inhibitors and provide Kivalues of 12.9 ± 4.6 mM and 7.9 ± 2.2 mM, respectively. Betaine 2 is a noncompetitive inhibitor with values for Kiiof 1.54 ± 0.31 mM and for Kisof 1.96 ± 1.36 mM. In each case enzyme activity was determined by measurement of (a) the 14C02liberated from the coupled decarboxylation of [l-14C]-2-oxoglutarate and (b) the 3H released into the aqueous medium from [2.3-3H]-γ-butyrobetaine added as substrate. None of the three cyclopropyl derivatives exhibited turnover, and preincubation of these substrate analogues with γ-butyrobetaine hydroxylase did not result in a time-dependent loss of activity greater than that of the controls. These results are discussed in terms of the conformation of the inhibitors relative to those of the substrate. © 1990, American Chemical Society. All rights reserved.