TY - JOUR
T1 - Capturing the Binuclear Copper State of Peptidylglycine Monooxygenase Using a Peptidyl-Homocysteine Lure
AU - Rush, Katherine W.
AU - Eastman, Karsten A.S.
AU - Welch, Evan F.
AU - Bandarian, Vahe
AU - Blackburn, Ninian J.
N1 - Publisher Copyright:
© 2024 American Chemical Society.
PY - 2024/2/28
Y1 - 2024/2/28
N2 - Peptidylglycine monooxygenase is a copper-dependent enzyme that catalyzes C-alpha hydroxylation of glycine extended pro-peptides, a critical post-translational step in peptide hormone processing. The canonical mechanism posits that dioxygen binds at the mononuclear M-center to generate a Cu(II)-superoxo species capable of H atom abstraction from the peptidyl substrate, followed by long-range electron tunneling from the CuH center. Recent crystallographic and biochemical data have challenged this mechanism, suggesting instead that an “open-to-closed” transition brings the copper centers closer, allowing reactivity within a binuclear intermediate. Here we present the first direct observation of an enzyme-bound binuclear copper species, captured by the use of an Ala-Ala-Phe-hCys inhibitor complex. This molecule reacts with the fully reduced enzyme to form a thiolate-bridged binuclear species characterized by EXAFS of the WT and its M314H variant and with the oxidized enzyme to form a novel mixed valence entity characterized by UV/vis and EPR. Mechanistic implications are discussed.
AB - Peptidylglycine monooxygenase is a copper-dependent enzyme that catalyzes C-alpha hydroxylation of glycine extended pro-peptides, a critical post-translational step in peptide hormone processing. The canonical mechanism posits that dioxygen binds at the mononuclear M-center to generate a Cu(II)-superoxo species capable of H atom abstraction from the peptidyl substrate, followed by long-range electron tunneling from the CuH center. Recent crystallographic and biochemical data have challenged this mechanism, suggesting instead that an “open-to-closed” transition brings the copper centers closer, allowing reactivity within a binuclear intermediate. Here we present the first direct observation of an enzyme-bound binuclear copper species, captured by the use of an Ala-Ala-Phe-hCys inhibitor complex. This molecule reacts with the fully reduced enzyme to form a thiolate-bridged binuclear species characterized by EXAFS of the WT and its M314H variant and with the oxidized enzyme to form a novel mixed valence entity characterized by UV/vis and EPR. Mechanistic implications are discussed.
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U2 - 10.1021/jacs.3c14705
DO - 10.1021/jacs.3c14705
M3 - Article
C2 - 38363651
AN - SCOPUS:85186078002
SN - 0002-7863
VL - 146
SP - 5074
EP - 5080
JO - Journal of the American Chemical Society
JF - Journal of the American Chemical Society
IS - 8
ER -