Phentermine inhibition of recombinant human liver monoamine oxidases A and B.

Recent studies with rat tissue preparations have suggested that the anorectic drug phentermine inhibits serotonin degradation by inhibition of monoamine oxidase (MAO) A with a K(I) value of 85-88 microM, a potency suggested to be similar to that of other reversible MAO inhibitors (Ulus et al., Biochem Pharmacol 2000;59:1611-21). Since there are known differences between rats and humans in substrate and inhibitor specificities of MAOs, the interactions of phentermine with recombinant human purified preparations of MAO A and MAO B were determined. Human MAO A was competitively inhibited by phentermine with a K(I) value of 498+/-60 microM, a value approximately 6-fold weaker than that observed for the rat enzyme. Phentermine was also observed to be a competitive inhibitor of recombinant human liver MAO B with a K(I) value of 375+/-42 microM, a value similar to that observed with the rat enzyme (310-416 microM). In contrast to the behavior with rat tissue preparations, no slow time-dependent behavior was observed for phentermine inhibition of purified soluble human MAO preparations. Difference absorption spectral studies showed similar perturbations of the covalent FAD moieties of both human MAO A and MAO B, which suggests a similar mode of binding in both enzymes. These data suggest that phentermine inhibition of human MAO A (or of MAO B) is too weak to be of pharmacological relevance.

Mechanism of time-dependent inhibition of polypeptide deformylase by actinonin.

Polypeptide deformylase (PDF) is an essential bacterial metalloenzyme responsible for the removal of the N-formyl group from the N-terminal methionine of nascent polypeptides. Inhibition of bacterial PDF enzymes by actinonin, a naturally occurring antibacterial agent, has been characterized using steady-state and transient kinetic methods. Slow binding of actinonin to these enzymes is observed under steady-state conditions. Progress curve analysis is consistent with a two-step binding mechanism, in which tightening of the initial encounter complex (EI) results in a final complex (EI*) with an extremely slow, but observable, off-rate (t(1/2) for inhibitor dissociation >or=0.77 days). Stopped-flow measurement of PDF fluorescence confirms formation of EI and provides a direct measurement of the association rate. Rapid dilution studies establish that the potency of actinonin is enhanced by more than 2000-fold upon tightening of EI to form EI*, from K(i) = 530 nM (EI) to Ki*