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 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.
Bakke
Mikio
M
Research and Development Division, Kikkoman Corporation, 399 Noda, Noda City, Chiba 278-0037, Japan. mbakke@mail.kikkoman.co.jp
Kajiyama
Naoki
N
D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D002850:Chromatography, Gel; D003605:D-Amino-Acid Oxidase / Q000737:chemistry* / Q000378:metabolism; D003911:Dextrans / Q000737:chemistry*; D004795:Enzyme Stability; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000201:enzymology; D007700:Kinetics; D010084:Oxidation-Reduction; D011485:Protein Binding; D013379:Substrate Specificity; D013552:Swine; D013696:Temperature
D000596:Amino Acids; D003911:Dextrans; D005182:Flavin-Adenine Dinucleotide; D003605:D-Amino-Acid Oxidase