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Identification and role of ionizing functional groups at the active center of Rhodotorula gracilis D ‐amino acid oxidase
Author(s) -
Pollegioni Loredano,
Harris Christopher M.,
Molla Gianluca,
Pilone Mirella S.,
Ghisla Sandro
Publication year - 2001
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(01)02983-0
Subject(s) - d amino acid oxidase , oxidase test , chemistry , stereochemistry , active center , ligand (biochemistry) , substrate (aquarium) , rhodotorula , flavoprotein , biochemistry , active site , alanine , binding site , residue (chemistry) , tyrosine , enzyme , amino acid , receptor , biology , yeast , ecology
D ‐Amino acid oxidase (DAAO) is a flavoprotein oxidase that catalyzes the oxidation of amino acids and produces ketoacids and H 2 O 2 . The rate of product release from reduced DAAO from Rhodotorula gracilis is pH dependent and reflects a p K a of ∼9.3. Binding of benzoate and 3,3,3‐trifluoro‐ D ‐alanine to wild‐type and Y238F–DAAO is also pH dependent (p K a =9.8±0.1 and 9.05±0.1, respectively for benzoate binding). However, binding of benzoate to Y223F–DAAO is pH independent, indicating the p K a is due to Y223–OH. This latter residue is thus involved in substrate binding, and probably is the group that governs product release. In contrast to this, the second active site tyrosine, Y238, has little influence on ligand binding.