Improvement in thermal stability and substrate binding of pig kidney D-amino acid oxidase by chemical modification

Chemical modification was evaluated to stabilize pig kidney D-amino acid oxidase (pkDAAO), which is required for analytical determination of D-amino acids. Optimization of modification conditions was performed to obtain high recovery yield and stability, and chemical modification at 30 degrees C for...

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Bibliographic Details
Published in:Applied biochemistry and biotechnology 2004-03, Vol.112 (3), p.123-132
Main Authors: Bakke, Mikio, Kajiyama, Naoki
Format: Article
Language:English
Subjects:
Amino acids
Amino Acids - metabolism
Animals
Biochemistry
Chromatography, Gel
D-Amino-Acid Oxidase - chemistry
D-Amino-Acid Oxidase - metabolism
Dextrans - chemistry
Enzyme Stability
Enzymes
Flavin-Adenine Dinucleotide - metabolism
Hogs
Hot Temperature
Hydrogen-Ion Concentration
Kidney - enzymology
Kidneys
Kinetics
Oxidation-Reduction
Protein Binding
Studies
Substrate Specificity
Swine
Temperature
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Summary:Chemical modification was evaluated to stabilize pig kidney D-amino acid oxidase (pkDAAO), which is required for analytical determination of D-amino acids. Optimization of modification conditions was performed to obtain high recovery yield and stability, and chemical modification at 30 degrees C for 12 h with a highly concentrated enzyme solution gave dextran-conjugated pkDAAO with a 70% yield of activity. pkDAAO was stable at less than 55 degrees C at pH 6.0, while the conjugated enzyme was stable even at 70 degrees C. In addition, the conjugated enzyme showed decreased Km values for D-amino acids. Because of these outstanding characteristics, this new material is expected to be available for use as a liquid assay reagent.
ISSN:0273-2289
0273-2289
1559-0291
DOI:10.1385/abab:112:3:123