| Texto completo | |
| Autor(es): |
Peskin, Alexander V.
;
Meotti, Flavia C.
;
Magon, Nicholas J.
;
de Souza, Luiz F.
;
Salvador, Armindo
;
Winterbourn, Christine C.
Número total de Autores: 6
|
| Tipo de documento: | Artigo Científico |
| Fonte: | Journal of Biological Chemistry; v. 301, n. 5, p. 15-pg., 2025-05-01. |
| Resumo | |
Peroxiredoxin 2 (Prdx2) undergoes ready glutathionylation, and glutaredoxin-catalyzed deglutathionylation provides an alternative mechanism to thioredoxin/thioredoxin reductase for recycling the reduced protein (Peskin et al. JBC 216, 3053, 2016). To elucidate the mechanism of glutathionylation, we have carried out kinetic studies using stopped flow and SDS PAGE plus product analysis by mass spectrometry. Kinetic modeling shows a mechanism in which exchange of Prdx2 disulfide with physiological concentrations of GSH occurs over seconds to minutes, initially at one active site to produce glutathionylated dimers linked by one disulfide. Exchange with GSH yields glutathionylation at both the peroxidatic (CP) and resolving cysteines (CR), the former predominating. Rate constants of 1.5 M-1s-1 and 0.021 s-1 were determined for exchange-mediated glutathionylation and deglutathionylation. Similar exchange reactions subsequently occur at the second active site. The rate of reaction of the CP sulfenic acid of WT Prdx2 with GSH (k = 10 M-1s-1) is 8 to 30 fold slower than when CR is mutated to Ser, Trp, or Asp and this reaction cannot effectively compete with intramolecular condensation. Consequently, when H2O2 reacts with reduced Prdx2 in the presence of GSH, the initial product is predominately the Prdx disulfide and glutathionylation subsequently occurs by exchange. However, glutathionylation of CR in the presence of H2O2 facilitates condensation of CP sulfenic acid with GSH to give diglutathionylated products and suppresses hyper-oxidation. This displaces equilibria and accelerates the conversion of Prdx2 to monomeric species. These results have implications for understanding the mechanism of relays between Prdx2 and other thiol proteins. (AU) | |
| Processo FAPESP: | 18/14898-2 - Processos redox na inflamação e o seu papel sobre doenças inflamatórias |
| Beneficiário: | Flavia Carla Meotti |
| Modalidade de apoio: | Auxílio à Pesquisa - Jovens Pesquisadores - Fase 2 |
| Processo FAPESP: | 13/07937-8 - Redoxoma |
| Beneficiário: | Ohara Augusto |
| Modalidade de apoio: | Auxílio à Pesquisa - Centros de Pesquisa, Inovação e Difusão - CEPIDs |
| Processo FAPESP: | 15/10411-3 - EMU concedido no processo 2013/08166-5: stopped-flow |
| Beneficiário: | Iolanda Midea Cuccovia |
| Modalidade de apoio: | Auxílio à Pesquisa - Programa Equipamentos Multiusuários |