Open AccessSite‐directed mutagenesis of potential catalytic residues in 1 H ‐3‐hydroxy‐4‐oxoquinoline 2,4‐dioxygenase, and hypothesis on the catalytic mechanism of 2,4‐dioxygenolytic ring cleavage
Author(s) -
Fischer Frank,
Fetzner Susanne
Publication year - 2000
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2000.tb09256.x
Subject(s) - dioxygenase , mutagenesis , cleavage (geology) , catalysis , chemistry , site directed mutagenesis , stereochemistry , ring (chemistry) , biochemistry , biology , mutation , gene , mutant , organic chemistry , paleontology , fracture (geology)
1 H ‐3‐Hydroxy‐4‐oxoquinoline 2,4‐dioxygenase (Qdo) is a cofactor‐free dioxygenase proposed to belong to the α/β hydrolase fold superfamily of enzymes. α/β Hydrolases contain a highly conserved catalytic triad (nucleophile–acidic residue–histidine). We previously identified a corresponding catalytically essential histidine residue in Qdo. However, as shown by amino acid replacements through site‐directed mutagenesis, nucleophilic and acidic residues of Qdo considered as possible triad residues were not absolutely required for activity. This suggests that Qdo does not contain the canonical catalytic triad of the α/β hydrolase fold enzymes. Some radical trapping agents affected the Qdo‐catalyzed reaction. A hypothetical mechanism of Qdo‐catalyzed dioxygenation of 1 H ‐3‐hydroxy‐4‐oxoquinoline is compared with the dioxygenation of FMNH 2 catalyzed by bacterial luciferase, which also uses a histidine residue as catalytic base.