Identification and role of ionizing functional groups at the active center of Rhodotorula gracilis D‐amino acid oxidase

FEBS Letters

Molla, Gianluca¹  Pilone, Mirella S.¹  Harris, Christopher M.¹  Ghisla, Sandro²  Pollegioni, Loredano¹ 

[1] Department of Structural and Functional Biology, University of Insubria, via J.H. Dunant 3, 21100 Varese, Italy;Faculty of Biology, University of Konstanz, P.O. Box 5560-M644, D-78434 Konstanz, Germany

关键词: D-Amino acid oxidase;    Flavoprotein;    pH effect;    Ligand binding;    Ionization;    Catalytic mechanism;    RgDAAO;    Rhodotorula gracilis D-amino acid oxidase;    pkDAAO;    pig kidney D-amino acid oxidase;    CF3-alanine;    3;    3;    3-trifluoro-alanine;    Bz;    benzoate;   

DOI  :  10.1016/S0014-5793(01)02983-0

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

D-Amino acid oxidase (DAAO) is a flavoprotein oxidase that catalyzes the oxidation of amino acids and produces ketoacids and H₂O₂. The rate of product release from reduced DAAO from Rhodotorula gracilis is pH dependent and reflects a pK _a of ∼9.3. Binding of benzoate and 3,3,3-trifluoro-D-alanine to wild-type and Y238F–DAAO is also pH dependent (pK _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 pK _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.

【 授权许可】

Unknown