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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Intra-dimer cooperativity between the active site cysteines during the oxidation of peroxiredoxin 2

Full text
Author(s):
Peskin, Alexander V. [1] ; Meotti, Flavia C. [2] ; Souza, Luiz F. de [2] ; Anderson, Robert F. [3] ; Winterbourn, Christine C. [1] ; Salvador, Armindo [4, 5, 6, 7]
Total Authors: 6
Affiliation:
[1] Univ Otago Christchurch, Dept Pathol & Biomed Sci, Ctr Free Radical Res, Christchurch - New Zealand
[2] Univ Sao Paulo, Dept Biochem, Chem Inst, Sao Paulo, SP - Brazil
[3] Univ Auckland, Sch Chem Sci, Auckland - New Zealand
[4] Univ Coimbra, Coimbra - Portugal
[5] Univ Coimbra, CNC Ctr Neurosci Cell Biol, Coimbra - Portugal
[6] CQC, Dept Chem, Coimbra - Portugal
[7] Univ Coimbra, Inst Interdisciplinary Res, Coimbra - Portugal
Total Affiliations: 7
Document type: Journal article
Source: Free Radical Biology and Medicine; v. 158, p. 115-125, OCT 2020.
Web of Science Citations: 0
Abstract

Peroxiredoxin 2 (Prdx2) and other typical 2-Cys Prdxs function as homodimers in which hydrogen peroxide oxidizes each active site cysteine to a sulfenic acid which then condenses with the resolving cysteine on the alternate chain. Previous kinetic studies have considered both sites as equally reactive. Here we have studied Prdx2 using a combination of non-reducing SDS-PAGE to separate reduced monomers and dimers with one and two disulfide bonds, and stopped flow analysis of tryptophan fluorescence, to investigate whether there is co-operativity between the sites. We have observed positive cooperativity when H2O2 is added as a bolus and oxidation of the second site occurs while the first site is present as a sulfenic acid. Modelling of this reaction showed that the second site reacts 2.2 +/- 0.1 times faster. In contrast, when H2O2 was generated slowly and the first active site condensed to a disulfide before the second site reacted, no cooperativity was evident. Conversion of the sulfenic acid to the disulfide showed negative cooperativity, with modelling of the exponential rise in tryptophan fluorescence yielding a rate constant of 0.75 +/- 0.08 s(-1) when the alternate active site was present as a sulfenic acid and 2.29 +/- 0.08-fold lower when it was a disulfide. No difference in the rate of hyperoxidation at the two sites was detected. Our findings imply that oxidation of one active site affects the conformation of the second site and influences which intermediate forms of the protein are favored under different cellular conditions. (AU)

FAPESP's process: 13/07937-8 - Redoxome - Redox Processes in Biomedicine
Grantee:Ohara Augusto
Support Opportunities: Research Grants - Research, Innovation and Dissemination Centers - RIDC
FAPESP's process: 18/14898-2 - Investigations of the redox processes in inflammatory response and associated pathologies
Grantee:Flavia Carla Meotti
Support Opportunities: Research Grants - Young Investigators Grants - Phase 2
FAPESP's process: 17/12312-8 - Role of Peroxiredoxins in the differentiation of myeloid leukemia cells and neutrophil function
Grantee:Luiz Felipe de Souza
Support Opportunities: Scholarships in Brazil - Post-Doctoral