Phosphonodipeptides containing (2-aminoethyl)phosphonic acid (ciliatine): transition state analogue inhibitors of carboxypeptidase A

Abstract

The phosphonodipeptides having a structure of Cbz-NH(CH 2) 2PO(O −Li +)-NHCHRCO 2 −Li + (Cbz = carbobenzyloxyl; amino acid residues Gly, l-Ala and l-Phe) were found to inhibit competitively the hydrolysis of Cbz-Gly-Gly- lPhe-OH by carboxypeptidase A. The compound having an l-phenylalanine residue was especially effective, giving K i values of about 10 −8 and 10 −9 M at pH 7.5 and 6.0 (25°C), respectively. The kinetics suggested that (i) an ionized P-O − group of the inhibitors with a tetrahedral phosphorus atom forms a coordinating bond with a zinc ion of the enzyme and (ii) the amino acid side-chains of the inhibitors are accommodated in the hydrophoic pocket of the enzyme. The inhibitory activity of the phosphonodipeptides was explained by a transition state analog mechanism.