2. Academic Validation
  3. Selective inactivation of the hydroxylase activity of bifunctional rat peptidylglycine alpha-amidating enzyme

Selective inactivation of the hydroxylase activity of bifunctional rat peptidylglycine alpha-amidating enzyme

  • Arch Biochem Biophys. 1992 May 1;294(2):594-602. doi: 10.1016/0003-9861(92)90730-k.
D J Merkler 1,  R Kulathila,  P P Tamburini,  S D Young
Affiliations

Affiliation

  • 1 Analytical Protein and Organic Chemistry Group, Unigene Laboratories, Inc., Fairfield, New Jersey 07004.
PMID: 1567215 DOI: 10.1016/0003-9861(92)90730-k
Abstract

Conversion of dansyl-Tyr-Val-Gly to dansyl-Tyr-Val-NH2 by recombinant type A rat 75-kDa peptidylglycine alpha-amidating enzyme (alpha-AE) is inactivated by ascorbate, dehydroascorbate, and hydrogen peroxide in a time- and concentration-dependent manner. Both ascorbate- and dehydroascorbate-mediated inactivation are saturable with apparent kinact/Kinact values of 1.7 and 0.23 s-1 M-1, respectively. Hydrogen peroxide-mediated inactivation is not saturable with a second-order rate constant of 50 s-1 M-1. Peptidyl-Gly substrates, EDTA, and H2O2 scavengers protect against ascorbate-mediated inactivation while EDTA and semidehydroascorbate scavengers protect against dehydroascorbate-mediated inactivation. Under similar conditions, ascorbate, dehydroascorbate, and H2O2 have no effect on the alpha-AE-catalyzed conversion of dansyl-Tyr-Val-alpha-hydroxyglycine to dansyl-Tyr-Val-NH2 which is consistent with the hypothesis that the 75-kDa enzyme consists of distinct peptidyl-Gly hydroxylase and peptidyl-alpha-hydroxyglycine lyase active sites.

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