The thioredoxin redox potential and redox charge are surrogate measures for flux in the thioredoxin system.

Abstract The thioredoxin system plays a central role in intracellular redox regulation and its dysregulation is associated with a number of pathologies. However, the connectivity within this system poses a significant challenge for quantification and consequently several disparate measures have been used to characterize the system. For in vitro studies, the thioredoxin system flux has been measured by NADPH oxidation while the thioredoxin redox state has been used to estimate the activity of the system in vivo. The connection between these measures has been obscure although substrate saturation in the thioredoxin system results from the saturation of the thioredoxin redox cycle. We used computational modeling and in vitro kinetic assays to clarify the relationship between flux and the current in vivo measures of the thioredoxin system together with a novel measure, the thioredoxin redox charge (reduced thioredoxin/total thioredoxin). Our results revealed that the thioredoxin redox potential and redox charge closely tracked flux perturbations showing that these indices could be used as surrogate measures of the flux in vivo and, provide a mechanistic explanation for the previously observed correlations between thioredoxin oxidation and certain pathologies. While we found no significant difference in the linear correlations obtained for the thioredoxin redox potential and redox charge with the flux, the redox charge may be preferred because it is bounded between zero and one and can be determined over a wider range of conditions allowing for quantitative flux comparisons between cell types and conditions.