Protein Thiol‐Based Probing of the Cellular and Metabolic Origins of Brain Oxidative Stress

Abstract

Improvement in this laboratory of a previously‐described method, exploiting the high affinity of vicinal thiols for phenylarsine oxide (PAO), has led to the development of a procedure for the efficient capture of proteins containing vicinal thiols that have been oxidized, in tissues, to disulfide bonds. Using this enhanced PAO‐affinity method, we have found that about 50% of the glycolytic enzyme triosephosphate isomerase (TPI) contains oxidized vicinal thiols, amidst a very low oxidation of total protein vicinal thiols, in brains from adult rats. Results of preliminary experiments employing a glutathione redox buffer argue that the vicinal thiols of TPI do not have an unusually high propensity for oxidation and support the view presented here that these thiols are highly oxidized in the brains under study because TPI is enriched in a population of cells that is experiencing substantial oxidative stress. We hypothesize that the observed TPI‐associated oxidative stress may result from increased rates of glucose metabolism by glycolysis at the expense of the antioxidant‐generating pentose phosphate pathway in TPI‐enriched cells. Our findings support the notion that protein thiols may serve as intrinsic sensors of the redox environments in which they reside and that the thiol redox states of cell type‐specific and metabolic state‐associated proteins may shed new light on the origins and, possibly, the pathological triggers of oxidative stress in complex tissues.