The hydrogen peroxide reactivity of peptidylglycine monooxygenase supports a Cu(II)-superoxo catalytic intermediate

Andrew T. Bauman, Erik T. Yukl, Katsiaryna Alkevich, Ashley L. McCormack, Ninian Blackburn

Research output: Contribution to journalArticle

43 Citations (Scopus)

Abstract

We have investigated the reaction of peptidylglycine monooxygenase with hydrogen peroxide to determine whether Cu(II)-peroxo is a likely intermediate. When the oxidized enzyme was reacted with the dansyl-YVG substrate and H 2O2, the α-hydroxyglycine product was formed. The reaction was catalytic and did not require the presence of additional reductant. When 18O-labeled H2O2 was reacted with peptidylglycine monooxygenase and substrate anaerobically, oxygen in the product was labeled with 18O and must therefore be derived from H 2O2. However, when the reaction was carried out with H2 16O2 in the presence of 18O 2, 60% of the product contained the 18O label. Therefore, the reaction must proceed via an intermediate that can react directly with dioxygen and thus scramble the label. Under strictly anaerobic conditions (in the presence of glucose and glucose oxidase, where no oxygen was released into the medium from nonenzymatic peroxide decomposition), product formation and peroxide consumption were tightly coupled, and the rate of product formation was identical to that measured under aerobic conditions. Peroxide reactivity was eliminated by a mutation at the CuH center, which should not be involved in the peroxide shunt. Our data lend support to recent proposals that Cu(II)-superoxide is the active species.

Original languageEnglish (US)
Pages (from-to)4190-4198
Number of pages9
JournalJournal of Biological Chemistry
Volume281
Issue number7
DOIs
StatePublished - Feb 17 2006

Fingerprint

Peroxides
Hydrogen Peroxide
Oxygen
Labels
Glucose Oxidase
Reducing Agents
Substrates
Superoxides
Decomposition
Glucose
Mutation
peptidylglycine monooxygenase
Enzymes

ASJC Scopus subject areas

  • Biochemistry

Cite this

The hydrogen peroxide reactivity of peptidylglycine monooxygenase supports a Cu(II)-superoxo catalytic intermediate. / Bauman, Andrew T.; Yukl, Erik T.; Alkevich, Katsiaryna; McCormack, Ashley L.; Blackburn, Ninian.

In: Journal of Biological Chemistry, Vol. 281, No. 7, 17.02.2006, p. 4190-4198.

Research output: Contribution to journalArticle

Bauman, Andrew T. ; Yukl, Erik T. ; Alkevich, Katsiaryna ; McCormack, Ashley L. ; Blackburn, Ninian. / The hydrogen peroxide reactivity of peptidylglycine monooxygenase supports a Cu(II)-superoxo catalytic intermediate. In: Journal of Biological Chemistry. 2006 ; Vol. 281, No. 7. pp. 4190-4198.
@article{37fd4899223a4dd49f375519f8ecf753,
title = "The hydrogen peroxide reactivity of peptidylglycine monooxygenase supports a Cu(II)-superoxo catalytic intermediate",
abstract = "We have investigated the reaction of peptidylglycine monooxygenase with hydrogen peroxide to determine whether Cu(II)-peroxo is a likely intermediate. When the oxidized enzyme was reacted with the dansyl-YVG substrate and H 2O2, the α-hydroxyglycine product was formed. The reaction was catalytic and did not require the presence of additional reductant. When 18O-labeled H2O2 was reacted with peptidylglycine monooxygenase and substrate anaerobically, oxygen in the product was labeled with 18O and must therefore be derived from H 2O2. However, when the reaction was carried out with H2 16O2 in the presence of 18O 2, 60{\%} of the product contained the 18O label. Therefore, the reaction must proceed via an intermediate that can react directly with dioxygen and thus scramble the label. Under strictly anaerobic conditions (in the presence of glucose and glucose oxidase, where no oxygen was released into the medium from nonenzymatic peroxide decomposition), product formation and peroxide consumption were tightly coupled, and the rate of product formation was identical to that measured under aerobic conditions. Peroxide reactivity was eliminated by a mutation at the CuH center, which should not be involved in the peroxide shunt. Our data lend support to recent proposals that Cu(II)-superoxide is the active species.",
author = "Bauman, {Andrew T.} and Yukl, {Erik T.} and Katsiaryna Alkevich and McCormack, {Ashley L.} and Ninian Blackburn",
year = "2006",
month = "2",
day = "17",
doi = "10.1074/jbc.M511199200",
language = "English (US)",
volume = "281",
pages = "4190--4198",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "7",

}

TY - JOUR

T1 - The hydrogen peroxide reactivity of peptidylglycine monooxygenase supports a Cu(II)-superoxo catalytic intermediate

AU - Bauman, Andrew T.

AU - Yukl, Erik T.

AU - Alkevich, Katsiaryna

AU - McCormack, Ashley L.

AU - Blackburn, Ninian

PY - 2006/2/17

Y1 - 2006/2/17

N2 - We have investigated the reaction of peptidylglycine monooxygenase with hydrogen peroxide to determine whether Cu(II)-peroxo is a likely intermediate. When the oxidized enzyme was reacted with the dansyl-YVG substrate and H 2O2, the α-hydroxyglycine product was formed. The reaction was catalytic and did not require the presence of additional reductant. When 18O-labeled H2O2 was reacted with peptidylglycine monooxygenase and substrate anaerobically, oxygen in the product was labeled with 18O and must therefore be derived from H 2O2. However, when the reaction was carried out with H2 16O2 in the presence of 18O 2, 60% of the product contained the 18O label. Therefore, the reaction must proceed via an intermediate that can react directly with dioxygen and thus scramble the label. Under strictly anaerobic conditions (in the presence of glucose and glucose oxidase, where no oxygen was released into the medium from nonenzymatic peroxide decomposition), product formation and peroxide consumption were tightly coupled, and the rate of product formation was identical to that measured under aerobic conditions. Peroxide reactivity was eliminated by a mutation at the CuH center, which should not be involved in the peroxide shunt. Our data lend support to recent proposals that Cu(II)-superoxide is the active species.

AB - We have investigated the reaction of peptidylglycine monooxygenase with hydrogen peroxide to determine whether Cu(II)-peroxo is a likely intermediate. When the oxidized enzyme was reacted with the dansyl-YVG substrate and H 2O2, the α-hydroxyglycine product was formed. The reaction was catalytic and did not require the presence of additional reductant. When 18O-labeled H2O2 was reacted with peptidylglycine monooxygenase and substrate anaerobically, oxygen in the product was labeled with 18O and must therefore be derived from H 2O2. However, when the reaction was carried out with H2 16O2 in the presence of 18O 2, 60% of the product contained the 18O label. Therefore, the reaction must proceed via an intermediate that can react directly with dioxygen and thus scramble the label. Under strictly anaerobic conditions (in the presence of glucose and glucose oxidase, where no oxygen was released into the medium from nonenzymatic peroxide decomposition), product formation and peroxide consumption were tightly coupled, and the rate of product formation was identical to that measured under aerobic conditions. Peroxide reactivity was eliminated by a mutation at the CuH center, which should not be involved in the peroxide shunt. Our data lend support to recent proposals that Cu(II)-superoxide is the active species.

UR - http://www.scopus.com/inward/record.url?scp=33645219172&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33645219172&partnerID=8YFLogxK

U2 - 10.1074/jbc.M511199200

DO - 10.1074/jbc.M511199200

M3 - Article

VL - 281

SP - 4190

EP - 4198

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 7

ER -