Pre-Steady-State Reactivity of Peptidylglycine Monooxygenase Implicates Ascorbate in Substrate Triggering of the Active Conformer.

ABSTRACT: Peptidylglycine monooxygenase (PHM) is essential for the posttranslational amidation of neuroendocrine peptides. An important aspect of the PHM mechanism is the complete coupling of oxygen reduction to substrate hydroxylation, which implies no oxygen reactivity of the fully reduced enzyme in the absence of peptidyl substrates. As part of studies aimed at investigating this feature of the PHM mechanism, we explored pre-steady-state kinetics using chemical quench (CQ) and rapid freeze-quench (RFQ) studies of the fully reduced ascorbate-free PHM enzyme. First, we confirmed the absence of Cu(I)-enzyme oxidation by O₂ at catalytic rates in the absence of peptidyl substrate. Next, we investigated reactivity in the presence of the substrate dansyl-YVG. Surprisingly, when ascorbate-free

SUBMITTER: Welch EF 

PROVIDER: S-EPMC9064607 | biostudies-literature | 2022 Apr

REPOSITORIES: biostudies-literature