The rate constant describing slow-onset inhibition of yeast AMP deaminase by coformycin analogs is independent of inhibitor structure

Publisher Website
Google Scholar
Abstract
(R)- and (S)-2''-doexycoformycin, (R)-coformycin, and the corresponding 5''-monophosphates were compared as inhibitors of yeast AMP deaminase. The overall inhibition constants ranged from 4.2 mM for (S)-2''-deoxycoformycin to 10 pM for (R)-coformycin 5''-monophosphate, a difference of 3.8 .times. 108 in affinities. (R)-Coformycin, (R)-2''-deoxycoformycin 5''-monophosphate, and (R)-coformycin-5''-monophosphate exhibited both rapid and slow-onset inhibition. The S inhibitors and (R)-2''-deoxycoformycin exhibited classical competitive inhibition but no time-dependent onset of inhibition. The results indicate that the presence of the 2''-hydroxyl and 5''-phosphate and the R stereochemistry at the C-8 position of the diazepine ring are necessary for the optimum interaction of inhibitors with yeast AMP deaminase. This differs from the results for rabbit muscle AMP deaminase [Frieden C., Kurz, L. C., and Gilbert, H. R. (1980) Biochemistry 19, 5303-5309] and calf intestinal adenosine deaminase [Schramm, V. L., and Baker, D. C. (1985) Biochemistry 24, 641-646], in which a tetrahedral hydroxyl at C-8 in the R stereochemistry is sufficient for slow-onset inhibition with the coformycins. The results suggest that the transition state contains a tetrahedral carbon with the R configuration as a result of the direct attack of an oxygen nucleophile at C-6 of AMP. Slow-onset inhibition of yeast AMP deaminase is consistent with a mechanism in which the combination of E and I is rapidly reversible. For these inhibitors, Ki varied by a factor of 3 .times. 103, and the overall inhibition constant (Ki*) varied by a factor of 2 .times. 105. The rate k5, which induces slow-onset inhibition, was similar for the three slow-onset inhibitors while k6 varied by a factor of 650, suggesting that k5 is independent of inhibitor structure. The ability to act as a slow-onset inhibitor is a result of the ability of the inhibitor to stabilize the EI* complex. These findings argue against inhibitior-induced transition-state conformations for the slow-onset inhibitors and suggest that the enzyme attains the transition-state configuration at a low rate independent of the inhibitor structure.

This publication has 24 references indexed in Scilit: