Inhibition of rabbit lung angiotensin-converting enzyme by N(α)-[(S)-1-carboxy-3-phenylpropyl]L-alanyl-L-proline and N(α)-[(S)-1-carboxy-3-phenylpropyl]L-lysyl-L-proline

H. G. Bull, N. A. Thornberry, M. H.J. Cordes, A. A. Patchett

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86 Scopus citations

Abstract

Two novel peptide analogs, N(α)-[(S)-1-carboxy-3-phenylpropyl]L-alanyl-L-proline and the corresponding L-lysyl-L-proline derivative, have been demonstrated to be potent competitive inhibitors of purified rabbit lung angiotensin-converting enzyme: K(i) = 2 and 1 x 10-10 M, respectively, at pH 7.5, 25°C, and 0.3 M chloride ion. Second-order rate constants for addition of these inhibitors to enzyme under the same conditions are in the range 1-2 x 106 M-1 s-1; first-order rate constants for dissociation of the EI complexes are in the range 1-4 x 10-4 s-1. The association rate constants are similar to those measured for D-3-mercapto-2-methylpropanoyl-L-proline, captopril, but the dissociation rate constants are severalfold slower and account for the higher affinity of these inhibitors for the enzyme. The dissociation constant for the EI complex containing N(α)-[(S)-1-carboxy-3-phenylpropyl]L-alanyl-L-proline is pH-dependent, and reaches a minimum at approximately pH 6: K(i) = 4 ± 1 x 10-11 M. The pH dependence is consistent either with a model for which the protonation state of the secondary nitrogen atom in the inhibitor determines binding affinity, or one for which ionizations on the enzyme alone influence affinity for these inhibitors. The affinity of this inhibitor for the zinc-free apoenzyme is 2 x 104 times less than that for the holoenzyme. If considered as a 'collected product' inhibitor, N(α)-[(S)-1-carboxy-3-phenylprolyl]L-alanyl-L-proline appears to derive an additional factor of 375 M in its affinity for the enzyme compared to that of the two products of its hypothetical hydrolysis, a consequence of favorably entropy effects.

Original languageEnglish (US)
Pages (from-to)2952-2962
Number of pages11
JournalJournal of Biological Chemistry
Volume260
Issue number5
StatePublished - May 29 1985
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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