Characterization of a peptide α-amidation activity from rat anterior pituitary

C. C. Glembotski, B. A. Eipper, R. E. Mains

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52 Scopus citations

Abstract

A secretory granule associated enzymatic activity from the rat anterior pituitary that can convert the synthetic peptide D-Tyr-Val-Gly into D-Tyr-Val-NH2 is described. Due to the presence of inhibitory activity in the cytosol and rough endoplasmic reticulum/Golgi apparatus fractions, the α-amidation activity is barely detectable in crude rat anterior pituitary homogenates. The α-amidation activity was primarily soluble, displayed a pH optimum of about 7.0, and showed a requirement for molecular oxygen. The activity was stimulated 7-fold by the addition of optimal concentrations of copper. Of the cofactors tested, only reduced ascorbate produced a severalfold stimulation of activity. Studies with varied D-Tyr-Val-Gly or varied monoiodo-D-Tyr-Val-Gly demonstrate Michaelis Menten kinetics with indistinguishable kinetic constants for both peptides. Upon addition of ascorbate, the apparent K(m) and V(max) for the synthetic substrate, as estimated from Lineweaver-Burk and Eadie-Hofstee plots, increased by 30- and 60-fold, respectively. Several α-melanotropin- and γ-melanotropin-related peptides with COOH-terminal glycine residues were effective competitive inhibitors of the reaction while the corresponding α-amidated peptides were very poor inhibitors. The rat anterior pituitary α-amidation activity appears to be very similar to the α-amidation activity in rat intermediate and neural pituitary, and mouse anterior pituitary tumor cells.

Original languageEnglish (US)
Pages (from-to)6385-6392
Number of pages8
JournalJournal of Biological Chemistry
Volume259
Issue number10
StatePublished - 1984
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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