Encapsulation of Trigonopsis variabilis D-amino acid oxidase and fast comparison of the operational stabilities of free and immobilized preparations of the enzyme.
A one-step procedure of immobilizing soluble and aggregated preparations of D-amino acid oxidase from Trigonopsis variabilis (TvDAO) is reported where carrier-free enzyme was entrapped in semipermeable microcapsules produced from the polycation poly(methylene-co-guanidine) in combination with CaCl2 and the polyanions alginate and cellulose sulfate. The yield of immobilization, expressed as the fraction of original activity present in microcapsules, was approximately 52 +/- 5%. The effectiveness of the entrapped oxidase for O2-dependent conversion of D-methionine at 25 degrees C was 85 +/- 10% of the free enzyme preparation. Because continuous spectrophotometric assays are generally not well compatible with insoluble enzymes, we employed a dynamic method for the rapid in situ estimation of activity and relatedly, stability of free and encapsulated oxidases using on-line measurements of the concentration of dissolved O2. Integral and differential modes of data acquisition were utilized to examine cases of fast and slow inactivation of the enzyme, respectively. With a half-life of 60 h, encapsulated TvDAO was approximately 720-fold more stable than the free enzyme under conditions of bubble aeration at 25 degrees C. The soluble oxidase was stabilized by added FAD only at temperatures of 35 degrees C or greater.
Nahalka
Jozef
J
Research Centre Applied Biocatalysis, Petersgasse 14, A-8010 Graz, Austria.
Dib
Iskandar
I
Nidetzky
Bernd
B
D002214:Capsules / Q000737:chemistry; D003605:D-Amino-Acid Oxidase / Q000302:isolation & purification* / Q000378:metabolism; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000378:metabolism; D006207:Half-Life; D004718:Saccharomycetales / Q000201:enzymology*
D002214:Capsules; D004800:Enzymes, Immobilized; D003605:D-Amino-Acid Oxidase