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Substituting selenocysteine for active site cysteine 149 of phosphorylating glyceraldehyde 3‐phosphate dehydrogenase reveals a peroxidase activity
Author(s) -
Boschi-Muller Sandrine,
Muller Sabine,
Van Dorsselaer Alain,
Böck August,
Branlant Guy
Publication year - 1998
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(98)01377-5
Subject(s) - selenocysteine , peroxidase , chemistry , active site , glyceraldehyde 3 phosphate dehydrogenase , ternary complex , dehydrogenase , stereochemistry , substrate (aquarium) , binding site , cysteine , enzyme , nad+ kinase , biochemistry , biology , ecology
Replacing the essential Cys‐149 by a selenocysteine into the active site of phosphorylating glyceraldehyde 3‐phosphate dehydrogenase (GAPDH) from Bacillus stearothermophilus leads to a selenoGAPDH that mimics a selenoperoxidase activity. Saturation kinetics were observed with cumenyl and tert ‐butyl hydroperoxides, with a better catalytic efficiency for the aromatic compound. The enzymatic mechanism fits a sequential model where the formation of a ternary complex between the holoselenoenzyme, the 3‐carboxy 4‐nitrobenzenethiol used as the reductant and the hydroperoxide precedes product release. The fact that the selenoGAPDH is NAD‐saturated supports a binding of hydroperoxide and reductant in the substrate binding site. The catalytic efficiency is similar to selenosubtilisins but remains low compared to selenoglutathione peroxidase. This is discussed in relation to what is known from the X‐ray crystal structures of selenoglutathione peroxidase and GAPDHs.