Bifunctional peptidylglycine α-amidating enzyme requires 2 copper atoms for maximum activity

Raviraj Kulathila, Angelo P. Consalvo, Paul F. Fitzpatrick, John C. Freeman, Lynn M. Snyder, Joseph J. Villafranca, David J. Merkler

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56 Scopus citations


The conversion of C-terminal glycine-extended peptides to C-terminal α-amidated peptides occurs in two distinct reactions, both of which are catalyzed by bifunctional peptidylglycine α-amidating enzyme. The first step is the α-hydroxylation of the C-terminal glycine residue and the second step is the dealkylation of the α-hydroxyglycine-extended peptide to the α-amidated peptide and glyoxylate. We show that the bifunctional enzyme requires 1.9 ± 0.2 mol of copper/mol of enzyme for maximal dansyl-Tyr-Lys-Gly amidation activity under the conditions of high enzyme concentration (∼80 μM) required to measure initial rates for this poor substrate. The enzyme, as purified, contains a substoichiometric amount of copper and has only trace levels of amidation activity. Addition of exogenous Cu(II) ions stimulates amidation activity ∼3000-fold at the optimum copper stoichiometry and the enzyme is then inhibited by excess Cu(II). No stimulation of amidation activity is observed upon the addition of the following divalent metal ions: Mn(II), Fe(II), NI(II), Cd(II), and the oxovanadium cation, VO(II). The enzyme-catalyzed dealkylation of α-hydroxyhippuric acid to benzamide shows no dependence on copper, indicating that the copper dependence of the amidation reaction must be attributed to a copper dependence in peptide α-hydroxylation.

Original languageEnglish (US)
Pages (from-to)191-195
Number of pages5
JournalArchives of Biochemistry and Biophysics
Issue number1
StatePublished - May 1994

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology


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