[17] S-Glutathionylation of glyceraldehyde-3-phosphate dehydrogenase: Role of thiol oxidation and catalysis by glutaredoxin

This chapter discusses methods employed to delineate the mechanism of S-glutathionylation of purified GAPDH, focusing on the roles of direct oxidation of the protein, with subsequent interaction of GSSG and the influence of GRX on this process. The chapter illustrates the complexity residing in the regulation of reversible S-glutathionylation of proteins, such as GAPDH, which is reflected in the design of suitable experimental approaches designed to cope with the interaction of several redox-dependent factors. Clear interactions are demonstrated between oxidative modification of GAPDH and its subsequent S-glutathionylation. Similarly, a redox interaction between GSSG and GAPDH with Grx as the catalyst is described, suggesting that the Grx molecule may participate in catalytic S-glutathionylation in intact cells. Grx itself can readily undergo S-glutathionylation, indicating the potential for regulation of this catalyst of the reversible S-glutathionylation of other proteins.