Biochemical characterizations of ER-resident peroxidases, GPx7 and GPx8, reveal their distinct and regulated oxidative activities

Abstract

In the mammalian endoplasmic reticulum (ER), the diverse network comprising more than 20 members of the protein disulfide isomerase (PDI) family and more than five PDI oxidases has evolved to promote oxidative protein folding. While the canonical disulfide bond formation pathway constituted by Ero1 and PDI has been well studied so far, mechanistic and physiological bases of newly identified PDI oxidases, glutathione peroxidase-7 (GPx7) and -8 (GPx8), are only poorly understood. We here demonstrated that human GPx7 has much higher reactivity with H2O2 than human GPx8, leading to efficient PDI oxidation. GPx7 forms a catalytic tetrad at the redox active site to react with H2O2 efficiently and stabilize a resultantly generated sulfenylated species. While it was previously postulated that the GPx7 catalysis involved a highly reactive peroxidatic cysteine, a resolving cysteine was found to act to regulate the PDI oxidation activity of GPx7. The present study also revealed that GPx7 formed complexes preferentially with PDI and P5 in H2O2-treated cells. Altogether, human GPx7 functions as an H2O2-dependent PDI oxidase in cells whereas PDI oxidation may not be the central physiological role of human GPx8.