Synthesis and evaluation of an inhibitor of carboxypeptidase A with a Ki value in the femtomolar range

Abstract

Comparative studies among a series of tripeptide phosphonate inhibitors of the zinc peptidase carboxypeptidase A indicate that incorporation of the phosphonic acid analogue of valine at the P1 position results in significantly higher affinity than the glycine, alanine, or phenylalanine analogues. When applied to the tripeptide analogue Cbz-Phe-ValP-(O)Phe [ZFVP(O)F], determination of the inhibition constant Ki was complicated by the very slow rate of dissociation. The rate of exchange of [3H]ZFVP(O)F with enzyme-bound [14C]ZFVP(O)F was followed for periods of 3-4 months to measure dissociation rate constants in the range of (1.7-4.4) x 10(-9) s-1, corresponding to half-lives of 5-13 years. Although the on- and off-rate constants differ for different carboxypeptidase isozymes, their ratios, corresponding to the inhibition constants Ki, are consistently in the range of 10-27 fM. Both the inhibition constants and the dissociation rate constants appear to be the lowest values yet determined for an enzyme-small inhibitor interaction.

Methods for the synthesis of α-heterocyclic/heteroaryl-α-aminophosphonic acids and their esters

“…These features explain the large range of biological activities displayed by the members of this important class of compounds and the applications. [2][3][4][5] They have been found in areas ranging from medicine to agriculture, for example, as antibiotics, 6 enzyme inhibitors, 7 anticancer agents 8 and herbicides. 9,10 These biological properties mostly are associated with the tetrahedral structure of the phosphonyl group acting as a "transition-state" analogue.…”

Conformation of Sterically Hindered 4-Methyl-2-oxo-2-trityl-1,3,2-dioxaphosphorinane in the Solid State and the Solution

“…The 1,3,2-dioxaphosphorinane derivatives, an important class of organophosphorus heterocycles, are interesting compounds because of their biological activities, particularly in reference to the design of enzyme inhibitors [51][52][53][54][55][56][57][58]. As a part of organophosphorus compounds the 1,3,2-dioxaphosphorinanes are also very important in pesticide and medicinal science, owing to their wide biological activities and stereochemistry [59].…”

Cyclen-Containing Inhibitors of Carboxypeptidase A Synthesized in Search of Target-Selective Artificial Proteases

“…To reach the new equilibrium, EI complex should dissociate to E and I which can take long when K i is very low. 4,5 For the potent inhibitors examined in this study, the new equilibrium was attained within 1-2 hours as checked by assay with ClCPL. Dissociation of EI during readjustment of the equilibrium states demonstrates the reversible nature of the EI formation in the case of the Cyc-containing inhibitors.…”

“…4,5 The X-ray crystallographic study performed on the phosphonate inhibitors disclosed that the L-β-phenyllactic acid (PLA) moiety containing the hydroxyalkylphosphinyl group occupied the active site. 6 Thus, the hydroxyalkylphosphinyl ester (A 0 ) of PLA can be exploited as binding sites of the CPA-cleaving catalysts.…”