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Reduction of sulfenic acids by ascorbate in proteins, connecting thiol-dependent to alternative redox pathways.
Free Radic Biol Med. 2020 Jun 29; 156:207-216.FR

Abstract

Sulfenic acids are the primary product of thiol oxidation by hydrogen peroxide and other oxidants. Several aspects of sulfenic acid formation through thiol oxidation were established recently. In contrast, the reduction of sulfenic acids is still scarcely investigated. Here, we characterized the kinetics of the reduction of sulfenic acids by ascorbate in several proteins. Initially, we described the crystal structure of our model protein (Tsa2-C170S). There are other Tsa2 structures in distinct redox states in public databases and all of them are decamers, with the peroxidatic cysteine very accessible to reductants, convenient features to investigate kinetics. We determined that the reaction between Tsa2-C170S-Cys-SOH and ascorbate proceeded with a rate constant of 1.40 ± 0.08 × 103 M-1 s-1 through a competition assay developed here, employing 2,6-dichlorophenol-indophenol (DCPIP). A series of peroxiredoxin enzymes (Prx6 sub family) were also analyzed by this competition assay and we observed that the reduction of sulfenic acids by ascorbate was in the 0.4-2.2 × 103 M-1 s-1 range. We also evaluated the same reaction on glyceraldehyde 3-phosphate dehydrogenase and papain, as the reduction of their sulfenic acids by ascorbate were reported previously. Once again, the rate constants are in the 0.4-2.2 × 103 M-1 s-1 range. We also analyzed the reduction of Tsa2-C170S-SOH by ascorbate by a second, independent method, following hydrogen peroxide reduction through a specific electrode (ISO-HPO-2, World Precision Instruments) and employing a bi-substrate, steady state approach. The kcat/KMAsc was 7.4 ± 0.07 × 103 M-1 s-1, which was in the same order of magnitude as the value obtained by the DCPIP competition assay. In conclusion, our data indicates that reduction of sulfenic acid in various proteins proceed at moderate rate and probably this reaction is more relevant in biological systems where ascorbate concentrations are high.

Authors+Show Affiliations

Departamento de Genética e Biologia Evolutiva, Instituto de Biociências, Universidade de São Paulo, 05508-090, São Paulo, Brazil.Laboratorio de Fisicoquímica Biológica, Instituto de Química Biológica, Facultad de Ciencias, Universidad de La República, Iguá 4225, Montevideo, 11400, Uruguay; Centro de Investigaciones Biomédicas (CEINBIO), Universidad de La República, Montevideo, Uruguay.Departamento de Genética e Biologia Evolutiva, Instituto de Biociências, Universidade de São Paulo, 05508-090, São Paulo, Brazil.Departamento de Genética e Biologia Evolutiva, Instituto de Biociências, Universidade de São Paulo, 05508-090, São Paulo, Brazil.Departamento de Genética e Biologia Evolutiva, Instituto de Biociências, Universidade de São Paulo, 05508-090, São Paulo, Brazil.Brazilian Biorenewables National Laboratory, National Center for Research in Energy and Materials, Campinas, Brazil.Brazilian Biorenewables National Laboratory, National Center for Research in Energy and Materials, Campinas, Brazil.Instituto de Biociências, UNESP, Campus Do Litoral Paulista, São Vicente, 11330-900, São Paulo, Brazil.Departamento de Genética e Biologia Evolutiva, Instituto de Biociências, Universidade de São Paulo, 05508-090, São Paulo, Brazil. Electronic address: nettoles@ib.usp.br.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

32615144

Citation

Anschau, Valesca, et al. "Reduction of Sulfenic Acids By Ascorbate in Proteins, Connecting Thiol-dependent to Alternative Redox Pathways." Free Radical Biology & Medicine, vol. 156, 2020, pp. 207-216.
Anschau V, Ferrer-Sueta G, Aleixo-Silva RL, et al. Reduction of sulfenic acids by ascorbate in proteins, connecting thiol-dependent to alternative redox pathways. Free Radic Biol Med. 2020;156:207-216.
Anschau, V., Ferrer-Sueta, G., Aleixo-Silva, R. L., Bannitz Fernandes, R., Tairum, C. A., Tonoli, C. C. C., Murakami, M. T., de Oliveira, M. A., & Netto, L. E. S. (2020). Reduction of sulfenic acids by ascorbate in proteins, connecting thiol-dependent to alternative redox pathways. Free Radical Biology & Medicine, 156, 207-216. https://doi.org/10.1016/j.freeradbiomed.2020.06.015
Anschau V, et al. Reduction of Sulfenic Acids By Ascorbate in Proteins, Connecting Thiol-dependent to Alternative Redox Pathways. Free Radic Biol Med. 2020 Jun 29;156:207-216. PubMed PMID: 32615144.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Reduction of sulfenic acids by ascorbate in proteins, connecting thiol-dependent to alternative redox pathways. AU - Anschau,Valesca, AU - Ferrer-Sueta,Gerardo, AU - Aleixo-Silva,Rogerio Luis, AU - Bannitz Fernandes,Renata, AU - Tairum,Carlos A, AU - Tonoli,Celisa Caldana Costa, AU - Murakami,Mario Tyago, AU - de Oliveira,Marcos Antonio, AU - Netto,Luis Eduardo Soares, Y1 - 2020/06/29/ PY - 2020/03/21/received PY - 2020/05/25/revised PY - 2020/06/06/accepted PY - 2020/7/3/pubmed PY - 2020/7/3/medline PY - 2020/7/3/entrez KW - Ascorbate KW - Peroxides KW - Peroxiredoxin KW - Sulfenic acid SP - 207 EP - 216 JF - Free radical biology & medicine JO - Free Radic. Biol. Med. VL - 156 N2 - Sulfenic acids are the primary product of thiol oxidation by hydrogen peroxide and other oxidants. Several aspects of sulfenic acid formation through thiol oxidation were established recently. In contrast, the reduction of sulfenic acids is still scarcely investigated. Here, we characterized the kinetics of the reduction of sulfenic acids by ascorbate in several proteins. Initially, we described the crystal structure of our model protein (Tsa2-C170S). There are other Tsa2 structures in distinct redox states in public databases and all of them are decamers, with the peroxidatic cysteine very accessible to reductants, convenient features to investigate kinetics. We determined that the reaction between Tsa2-C170S-Cys-SOH and ascorbate proceeded with a rate constant of 1.40 ± 0.08 × 103 M-1 s-1 through a competition assay developed here, employing 2,6-dichlorophenol-indophenol (DCPIP). A series of peroxiredoxin enzymes (Prx6 sub family) were also analyzed by this competition assay and we observed that the reduction of sulfenic acids by ascorbate was in the 0.4-2.2 × 103 M-1 s-1 range. We also evaluated the same reaction on glyceraldehyde 3-phosphate dehydrogenase and papain, as the reduction of their sulfenic acids by ascorbate were reported previously. Once again, the rate constants are in the 0.4-2.2 × 103 M-1 s-1 range. We also analyzed the reduction of Tsa2-C170S-SOH by ascorbate by a second, independent method, following hydrogen peroxide reduction through a specific electrode (ISO-HPO-2, World Precision Instruments) and employing a bi-substrate, steady state approach. The kcat/KMAsc was 7.4 ± 0.07 × 103 M-1 s-1, which was in the same order of magnitude as the value obtained by the DCPIP competition assay. In conclusion, our data indicates that reduction of sulfenic acid in various proteins proceed at moderate rate and probably this reaction is more relevant in biological systems where ascorbate concentrations are high. SN - 1873-4596 UR - https://www.unboundmedicine.com/medline/citation/32615144/Reduction_of_sulfenic_acids_by_ascorbate_in_proteins,_connecting_thiol-dependent_to_alternative_redox_pathways L2 - https://linkinghub.elsevier.com/retrieve/pii/S0891-5849(20)31112-6 DB - PRIME DP - Unbound Medicine ER -
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