Competitive oxidation of key pentose phosphate pathway enzymes modulates the fate of intermediates and NAPDH production

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dc.catalogadorjwg
dc.contributor.authorReyes Valenzuela Juan, Sebastián
dc.contributor.authorCortés Ríos, Javiera Alejandra
dc.contributor.authorFuentes-Lemus E.
dc.contributor.authorRodríguez Fernandez, María
dc.contributor.authorDavies M.J.
dc.contributor.authorLopéz Alarcón, Camilo
dc.date.accessioned2025-01-02T19:17:46Z
dc.date.available2025-01-02T19:17:46Z
dc.date.issued2024
dc.description.abstractThe oxidative phase of the pentose phosphate pathway (PPP) involving the enzymes glucose-6-phosphate dehydrogenase (G6PDH), 6-phosphogluconolactonase (6PGL), and 6-phosphogluconate dehydrogenase (6PGDH), is critical to NADPH generation within cells, with these enzymes catalyzing the conversion of glucose-6-phosphate (G6P) into ribulose-5-phosphate (Ribu5-P). We have previously studied peroxyl radical (ROO•) mediated oxidative inactivation of E. coli G6PDH, 6PGL, and 6PGDH. However, these data were obtained from experiments where each enzyme was independently exposed to ROO•, a condition not reflecting biological reality. In this work we investigated how NADPH production is modulated when these enzymes are jointly exposed to ROO•. Enzyme mixtures (1:1:1 ratio) were exposed to ROO• produced from thermolysis of 100 mM 2,2′-azobis(2-methylpropionamidine) dihydrochloride (AAPH). NADPH was quantified at 340 nm, and protein oxidation analyzed by liquid chromatography with mass spectrometric detection (LC-MS). The data obtained were rationalized using a mathematical model. The mixture of non-oxidized enzymes, G6P and NADP+ generated ∼175 μM NADPH. Computational simulations showed a constant decrease of G6P associated with NADPH formation, consistent with experimental data. When the enzyme mixture was exposed to AAPH (3 h, 37 °C), lower levels of NADPH were detected (∼100 μM) which also fitted with computational simulations. LC-MS analyses indicated modifications at Tyr, Trp, and Met residues but at lower concentrations than detected for the isolated enzymes. Quantification of NADPH generation showed that the pathway activity was not altered during the initial stages of the oxidations, consistent with a buffering role of G6PDH towards inactivation of the oxidative phase of the pathway.
dc.description.funderFondecyt
dc.description.funderNovo Nordisk Foundation
dc.description.funderFONDEQUIP
dc.fuente.origenORCID
dc.identifier.doi10.1016/j.freeradbiomed.2024.05.050
dc.identifier.issn18734596 08915849
dc.identifier.scopusidSCOPUS_ID:85197793234
dc.identifier.urihttps://repositorio.uc.cl/handle/11534/89463
dc.information.autorucEscuela de Química; Fuentes Lemus, Eduardo Felipe; 0000-0002-1465-8466; 186720
dc.information.autorucEscuela de Química; Reyes Valenzuela Juan Sebastian; S/I; 1268701
dc.information.autorucEscuela de Ingeniería; Cortes Rios Javiera Alejandr; S/I; 1092020
dc.information.autorucEscuela de Ingeniería; Rodriguez Fernandez Maria; 0003 1966 2920; 1031920
dc.language.isoen
dc.nota.accesocontenido completo
dc.pagina.final518
dc.pagina.inicio505
dc.publisherElsevier Inc.
dc.revistaFree Radical Biology and Medicine
dc.rightsacceso restringido
dc.subject6-phosphogluconolactonase
dc.subjectAAPH
dc.subjectGlucose-6-phosphate dehydrogenase
dc.subjectMathematical simulations
dc.subjectNADPH
dc.subjectPeroxyl radicals
dc.subjectProtein oxidation
dc.subject.ddc570
dc.subject.deweyBiologíaes_ES
dc.titleCompetitive oxidation of key pentose phosphate pathway enzymes modulates the fate of intermediates and NAPDH production
dc.typeartículo
dc.volumen222
sipa.codpersvinculados186720
sipa.codpersvinculados1268701
sipa.codpersvinculados1092020
sipa.codpersvinculados1031920
sipa.trazabilidadORCID;2024-12-23
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