| Role of the active site residues arginine-216 and arginine-237 in the substrate specificity of mammalian D-aspartate oxidase. | |
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MedLine Citation:
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PMID: 20567862 Owner: NLM Status: In-Process |
Abstract/OtherAbstract:
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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. |
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Authors:
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Masumi Katane; Yasuaki Saitoh; Kazuhiro Maeda; Toshihiko Hanai; Masae Sekine; Takemitsu Furuchi; Hiroshi Homma |
Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2010-06-22 |
Journal Detail:
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Title: Amino acids Volume: 40 ISSN: 1438-2199 ISO Abbreviation: Amino Acids Publication Date: 2011 Feb |
Date Detail:
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Created Date: 2011-01-13 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 9200312 Medline TA: Amino Acids Country: Austria |
Other Details:
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Languages: eng Pagination: 467-76 Citation Subset: IM |
Affiliation:
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Laboratory of Biomolecular Science, Department of Pharmaceutical Life Sciences, Kitasato University, 5-9-1 Shirokane, Minato-ku, Tokyo, 108-8641, Japan. |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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