| First Author | Katane M | Year | 2011 |
| Journal | Amino Acids | Volume | 40 |
| Issue | 2 | Pages | 467-76 |
| PubMed ID | 20567862 | Mgi Jnum | J:343404 |
| Mgi Id | MGI:7565911 | Doi | 10.1007/s00726-010-0658-4 |
| Citation | Katane M, et al. (2011) Role of the active site residues arginine-216 and arginine-237 in the substrate specificity of mammalian D-aspartate oxidase. Amino Acids 40(2):467-76 |
| abstractText | D-aspartate oxidase (DDO) and D-amino acid oxidase (DAO) are flavin adenine dinucleotide-containing flavoproteins that catalyze the oxidative deamination of D-amino acids. Unlike DAO, which acts on several neutral and basic D-amino acids, DDO is highly specific for acidic D-amino acids. Based on molecular modeling and simulated annealing docking analyses, a recombinant mouse DDO carrying two substitutions (Arg-216 to Leu and Arg-237 to Tyr) was generated (R216L-R237Y variant). This variant and two previously constructed single-point mutants of mouse DDO (R216L and R237Y variants) were characterized to investigate the role of Arg-216 and Arg-237 in the substrate specificity of mouse DDO. The R216L-R237Y and R216L variants acquired a broad specificity for several neutral and basic D-amino acids, and showed a considerable decrease in activity against acidic D-amino acids. The R237Y variant, however, did not show any additional specificity for neutral or basic D-amino acids and its activity against acidic D-amino acids was greatly reduced. The kinetic properties of these variants indicated that the Arg-216 residue is important for the catalytic activity and substrate specificity of mouse DDO. However, Arg-237 is, apparently, only marginally involved in substrate recognition, but is important for catalytic activity. Notably, the substrate specificity of the R216L-R237Y variant differed significantly from that of the R216L variant, suggesting that Arg-237 has subsidiary effects on substrate specificity. Additional experiments using several DDO and DAO inhibitors also suggested the involvement of Arg-216 in the substrate specificity and catalytic activity of mouse DDO and that Arg-237 is possibly involved in substrate recognition by this enzyme. Collectively, these results indicate that Arg-216 and Arg-237 play crucial and subsidiary role(s), respectively, in the substrate specificity of mouse DDO. |
