SELECTIVE INACTIVATION OF THE HYDROXYLASE-ACTIVITY OF BIFUNCTIONAL RAT PEPTIDYLGLYCINE ALPHA-AMIDATING ENZYME

Conversion of dansyl-TyrValGly to dansyl-TyrValNH2 by recombinant type A rat 75-kDa peptidylglycine α-amidating enzyme (α-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 α-AE-catalyzed conversion of dansyl-TyrValα-hydroxyglycine to dansyl-TyrValNH2 which is consistent with the hypothesis that the 75-kDa enzyme consists of distinct peptidyl-Gly hydroxylase and peptidyl-α-hydroxyglycine lyase active sites. © 1992.