The most recent research on D-amino acid oxidases and D-amino acid metabolism has revealed new, intriguing properties of flavoenzymes and enlighted novel biotechnological uses of this catalyst Concerning the in vivo function of the enzyme, new findings on the physiological role of D-amino acid oxidase point to a detoxifying function of the enzyme in metabolizing exogenous D-amino acids in animals A novel role in modulating the level of D-serine in brain has also been proposed for the enzyme At the molecular level, site-directed mutagenesis studies on the pig kidney D-amino acid oxidase and, more recently, on the enzyme from the yeast Rhodotorula gracilis indicated that the few conserved residues of the active site do not play a role in acid-base catalysis but rather are involved in substrate interactions The three-dimensional structure of the enzyme was recently determined from two different sources: at 25–30 A resolution for DAAO from pig kidney and at 12–18 A resolution for R gracilis The active site can be clearly depicted: the striking absence of essential residues acting in acid-base catalysis and the mode of substrate orientation into the active site, taken together with the results of free-energy correlation studies, clearly support a hydrid transfer type of mechanism in which the orbital steering between the substrate and the isoalloxazine atoms plays a crucial role during catalysis

D-Amino acid oxidase: new findings.

The coimmobilization of d-amino acid oxidase and catalase enables the quantitative transformation of d-amino acids (d-phenylalanine) into α-keto acids (phenylpyruvic acid)

Co-aggregation of Enzymes and Polyethyleneimine: A Simple Method To Prepare Stable and Immobilized Derivatives of Glutaryl Acylase

Biocatalysis driven by D-amino acid oxidase is a significant example of the commercial production of high value-added intermediates using enzyme-based technology. The results of the most recent research on this FAD-dependent catalyst are reported here. In particular, insight is given of how in the past few years the main industrial application of this enzyme, i.e. the stereospecific bioconversion of cephalosporin C to glutaryl-7-amino cephalosporanic acid in the two-step production of 7-amino cephalosporanic acid, has been implemented by improving its production and by engineering of the biocatalyst. The set-up and the optimization of different conditions for carrying out the process under different procedures have also been updated.

D-amino acid oxidase as an industrial biocatalyst

Xylanase from Bacillus pumilus strain MK001 was immobilized on different matrices following varied immobilization methods. Entrapment using gelatin (GE) (40.0%), physical adsorption on chitin (CH) (35.0%), ionic binding with Q-sepharose (Q-S) (45.0%), and covalent binding with HP-20 beads (42.0%) showed the maximum xylanase immobilization efficiency. The optimum pH of immobilized xylanase shifted up to 1.0 unit (pH 7.0) as compared to free enzyme (pH 6.0). The immobilized xylanase exhibited higher pH stability (up to 28.0%) in the alkaline pH range (7.0-10.0) as compared to free enzyme. Optimum temperature of immobilized xylanase was observed to be 8 degrees C higher (68.0 degrees C) than free enzyme (60.0 degrees C). The free xylanase retained 50.0% activity, whereas xylanase immobilized on HP-20, Q-S, CH, and GE retained 68.0, 64.0, 58.0, and 57.0% residual activity, respectively, after 3 h of incubation at 80.0 degrees C. The immobilized xylanase registered marginal increase and decrease in Km and Vmax values, respectively, as compared to free enzyme. The immobilized xylanase retained up to 70.0% of its initial hydrolysis activity after seven enzyme reaction cycles. The immobilized xylanase was found to produce higher levels of high-quality xylo-oligosaccharides from birchwood xylan, indicating its potential in the nutraceutical industry.

R. Tassinari

Papers

Immobilization of d-amino acid oxidase from different yeasts: Characterization and application in the deamination of cephalosporin C

Immobilization of glutaryl-7-ACA acylase on aminoalkylated polyacrylic supports