Copper ion / H2O2 oxidation of Cu/Zn-Superoxide dismutase: Implications for enzymatic activity and antioxidant action

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

Standard

Copper ion / H2O2 oxidation of Cu/Zn-Superoxide dismutase : Implications for enzymatic activity and antioxidant action. / Tiwari, Manish K.; Hägglund, P. M.; Møller, Ian Max; Davies, Michael J.; Bjerrum, Morten J.

In: Redox Biology, Vol. 26, 101262, 2019.

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

Harvard

APA

CBE

MLA

Vancouver

Author

Tiwari, Manish K. ; Hägglund, P. M. ; Møller, Ian Max ; Davies, Michael J. ; Bjerrum, Morten J. / Copper ion / H2O2 oxidation of Cu/Zn-Superoxide dismutase : Implications for enzymatic activity and antioxidant action. In: Redox Biology. 2019 ; Vol. 26.

Bibtex

@article{d32a402c651c46d7b2b3b029d8267111,
title = "Copper ion / H2O2 oxidation of Cu/Zn-Superoxide dismutase: Implications for enzymatic activity and antioxidant action",
abstract = "Copper ion-catalyzed oxidation of yeast SOD1 (ySOD1) was examined to determine early oxidative modifications, including oxidation of a crucial disulfide bond, and the structural and functional repercussions of these events. The study used distinct oxidative conditions: Cu2+/H2O2, Cu2+/H2O2/AscH− and Cu2+/H2O2/glucose. Capillary electrophoresis experiments and quantification of protein carbonyls indicate that ySOD1 is highly susceptible to oxidative modification and that changes can be detected within 0.1 min of the initiation of the reaction. Oxidation-induced structural perturbations, characterized by circular dichroism, revealed the formation of partially-unfolded ySOD1 species in a dose-dependent manner. Consistent with these structural changes, pyrogallol assay indicates a partial loss of enzymatic activity. ESI-MS analyses showed seven distinct oxidized ySOD1 species under mild oxidation within 0.1 min. LC/MS analysis after proteolytic digestion demonstrated that the copper-coordinating active site histidine residues, His47 and His49, were converted into 2-oxo-histidine. Furthermore, the Cu and Zn bridging residue, His64 is converted into aspartate/asparagine. Importantly, the disulfide-bond Cys58-Cys147 which is critical for the structural and functional integrity of ySOD1 was detected as being oxidized at Cys147. We propose, based on LC/MS analyses, that disulfide-bond oxidation occurs without disulfide bond cleavage. Modifications were also detected at Met85 and five surface-exposed Lys residues. Based on these data we propose that the Cys58-Cys147 bond may act as a sacrificial target for oxidants and protect ySOD1 from oxidative inactivation arising from exposure to Cu2+/H2O2 and auto-inactivation during extended enzymatic turnover.",
keywords = "Amyotrophic lateral sclerosis, Disulfide oxidation, Metal-ion catalyzed oxidation, Oxidative stress, Protein carbonyls, Superoxide dismutase",
author = "Tiwari, {Manish K.} and H{\"a}gglund, {P. M.} and M{\o}ller, {Ian Max} and Davies, {Michael J.} and Bjerrum, {Morten J.}",
year = "2019",
doi = "10.1016/j.redox.2019.101262",
language = "English",
volume = "26",
journal = "Redox Biology",
issn = "2213-2317",
publisher = "Elsevier BV",

}

RIS

TY - JOUR

T1 - Copper ion / H2O2 oxidation of Cu/Zn-Superoxide dismutase

T2 - Implications for enzymatic activity and antioxidant action

AU - Tiwari, Manish K.

AU - Hägglund, P. M.

AU - Møller, Ian Max

AU - Davies, Michael J.

AU - Bjerrum, Morten J.

PY - 2019

Y1 - 2019

N2 - Copper ion-catalyzed oxidation of yeast SOD1 (ySOD1) was examined to determine early oxidative modifications, including oxidation of a crucial disulfide bond, and the structural and functional repercussions of these events. The study used distinct oxidative conditions: Cu2+/H2O2, Cu2+/H2O2/AscH− and Cu2+/H2O2/glucose. Capillary electrophoresis experiments and quantification of protein carbonyls indicate that ySOD1 is highly susceptible to oxidative modification and that changes can be detected within 0.1 min of the initiation of the reaction. Oxidation-induced structural perturbations, characterized by circular dichroism, revealed the formation of partially-unfolded ySOD1 species in a dose-dependent manner. Consistent with these structural changes, pyrogallol assay indicates a partial loss of enzymatic activity. ESI-MS analyses showed seven distinct oxidized ySOD1 species under mild oxidation within 0.1 min. LC/MS analysis after proteolytic digestion demonstrated that the copper-coordinating active site histidine residues, His47 and His49, were converted into 2-oxo-histidine. Furthermore, the Cu and Zn bridging residue, His64 is converted into aspartate/asparagine. Importantly, the disulfide-bond Cys58-Cys147 which is critical for the structural and functional integrity of ySOD1 was detected as being oxidized at Cys147. We propose, based on LC/MS analyses, that disulfide-bond oxidation occurs without disulfide bond cleavage. Modifications were also detected at Met85 and five surface-exposed Lys residues. Based on these data we propose that the Cys58-Cys147 bond may act as a sacrificial target for oxidants and protect ySOD1 from oxidative inactivation arising from exposure to Cu2+/H2O2 and auto-inactivation during extended enzymatic turnover.

AB - Copper ion-catalyzed oxidation of yeast SOD1 (ySOD1) was examined to determine early oxidative modifications, including oxidation of a crucial disulfide bond, and the structural and functional repercussions of these events. The study used distinct oxidative conditions: Cu2+/H2O2, Cu2+/H2O2/AscH− and Cu2+/H2O2/glucose. Capillary electrophoresis experiments and quantification of protein carbonyls indicate that ySOD1 is highly susceptible to oxidative modification and that changes can be detected within 0.1 min of the initiation of the reaction. Oxidation-induced structural perturbations, characterized by circular dichroism, revealed the formation of partially-unfolded ySOD1 species in a dose-dependent manner. Consistent with these structural changes, pyrogallol assay indicates a partial loss of enzymatic activity. ESI-MS analyses showed seven distinct oxidized ySOD1 species under mild oxidation within 0.1 min. LC/MS analysis after proteolytic digestion demonstrated that the copper-coordinating active site histidine residues, His47 and His49, were converted into 2-oxo-histidine. Furthermore, the Cu and Zn bridging residue, His64 is converted into aspartate/asparagine. Importantly, the disulfide-bond Cys58-Cys147 which is critical for the structural and functional integrity of ySOD1 was detected as being oxidized at Cys147. We propose, based on LC/MS analyses, that disulfide-bond oxidation occurs without disulfide bond cleavage. Modifications were also detected at Met85 and five surface-exposed Lys residues. Based on these data we propose that the Cys58-Cys147 bond may act as a sacrificial target for oxidants and protect ySOD1 from oxidative inactivation arising from exposure to Cu2+/H2O2 and auto-inactivation during extended enzymatic turnover.

KW - Amyotrophic lateral sclerosis

KW - Disulfide oxidation

KW - Metal-ion catalyzed oxidation

KW - Oxidative stress

KW - Protein carbonyls

KW - Superoxide dismutase

UR - http://www.scopus.com/inward/record.url?scp=85068395969&partnerID=8YFLogxK

U2 - 10.1016/j.redox.2019.101262

DO - 10.1016/j.redox.2019.101262

M3 - Journal article

C2 - 31284117

AN - SCOPUS:85068395969

VL - 26

JO - Redox Biology

JF - Redox Biology

SN - 2213-2317

M1 - 101262

ER -