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UNIVERSIDADE DE SÃO PAULO INSTITUTO DE QUÍMICA Programa de Pós-Graduação em Ciências Biológicas (Bioquímica)
| Content Provider | Semantic Scholar |
|---|---|
| Author | Santos, Nayra Fernandes |
| Copyright Year | 2014 |
| Abstract | PATRICIO, E. S. Evaluation of the interaction of urate hydroperoxide with protein disulfide isomerase (PDI) in inflammatory processes. 2014. 99 p. M. Sc. Dissertation – Graduate Program in Biochemistry, Chemistry Institute, University of Sao Paulo, Sao Paulo. The oxidation of the uric acid (7,9-dihidro-1H-purine-2,6,8(3H)-trione) by myeloperoxidase (MPO) generates the urate radical. In inflammatory conditions the superoxide reacts with urate radical to form the urate hydroperoxide (HOOU). Taking into account the high amount of MPO, urate and superoxide in the atheroma plaque, it is likely that HOOU is being formed in this inflammatory environment. The HOOU is a strong oxidizing agent and can react with thiol groups from proteins, like the protein disulfide isomerase (PDI). As a consequence of its oxidation, PDI positively modulates NADPH oxidase (Nox) and increases superoxide production by neutrophils. Therefore, the formation of HOOU in the vascular sheet would contribute to tissue damage and would explain the positive correlation between hyperuricemia and the risk for cardiovascular disease. To investigate the interaction of HOOU with PDI, we performed the chemical synthesis of the compound by the Type I photooxidation, using UVA irradiation and riboflavin as a photosensitizer. Initially, we standardized the irradiation light and the time of the reaction that produced the highest income. The HOOU and its reduced product 5hydroxiisourate were separate, identified and characterized by liquid chromatography coupled to mass spectrometry (LC/MS). We also determine the molar extinction coefficient of HOOU at 308 nm (ɛ308 = 6537 ± 377 M -1 .cm -1 ). The half-life of the compound was 41 min at 22 o C. The HOOU selectively oxidized methionine and glutathione. The reaction of HOOU with glutathione did not form any stable adducts. Thus, all consumed glutathione generated glutathione disulfide. When incubated with PDI (10 μM), 70 and 100% of the total amount of HOOU (3 μM) was consumed within 30 and 120 seconds, respectively. Besides, the PDI (23 μM) had its thiol groups oxidized after incubation with 140 μM HOOU for 30 min at 22oC. The HOOU oxidized the cysteine residues from the catalytical sites of PDI with a second order rate constant of 1.5 ± 0.04 x 10 3 M 1 .s -1 . This result suggests a favorable interaction of HOOU with this protein in the biological system, as well as a possible modulatory role of HOOU on the PDI-Nox pathway. Interestingly, urate increased superoxide production and oxygen consumption by neutrophil-like cells (differentiated HL-60 cells). This effect could be mediated by the formation of HOOU during the neutrophil oxidative burst, followed by the oxidation of PDI, a positive regulation of Nox and an increase in superoxide production. |
| File Format | PDF HTM / HTML |
| Alternate Webpage(s) | https://teses.usp.br/teses/disponiveis/46/46131/tde-01102014-090745/publico/DissertCorrigidaNayraFernandesSantos.pdf |
| Language | English |
| Access Restriction | Open |
| Content Type | Text |
| Resource Type | Article |
