Reactivity of glutaredoxins 1, 2 and 3 from Escherichia coli and protein disulfide isomerase towards glutathionyl-mixed disulfides in ribonuclease A

We have examined the activity of protein disulfide isomerase (PDI) and glut aredoxin (Grx) 1, 2 and 3 from Escherichia coli to catalyze the cleavage of glutathionylated ribonuclease A (RNase-SG) by 1 mM GSH to yield reduced RN ase. Apparent K-m values for RNase-SG were similar, 2-10 mu M, for Grx 1, 3 and PDI but Grx 1 and Grx 3 showed 500-fold higher turnover numbers than P DI, The atypical Grx 2 also catalyzed deglutathionylation by GSH, but had h igher K-m and apparent turnover number values compared to the two classical Grx. Refolding of RNase in a glutathione redox buffer was catalyzed by PDI . However, it could be measured only after a characteristic lag phase that was shortened by all three E. coli Grxs in a concentration-dependent manner . A role of the glutaredoxin mechanism in the endoplasmic reticulum is sugg ested. (C) 1999 Federation of European Biochemical Societies.