Catalytic properties of D-amino acid oxidase in cephalosporin C bioconversion: a comparison between proteins from different sources.
Lacking an efficient process to produce 7-aminocephalosporanic acid from cephalosporin C in a single step, d-amino acid oxidase (DAAO) is of foremost importance in the industrial, two-step process used for this purpose. We report a detailed study on the catalytic properties of the three available DAAOs, namely, a mammalian DAAO and two others from yeast (Rhodotorula gracilis and Trigonopsis variabilis). In comparing the kinetic parameters determined for the three DAAOs, with both cephalosporin C and d-alanine as substrate, the catalytic efficiency of the two enzymes from microorganism is at least 2 orders of magnitude higher than that of pig kidney DAAO. Furthermore, the mammalian enzyme is more sensitive to product inhibition (from hydrogen peroxide and glutaryl-7-aminocephalosporanic acid). Therefore, enzymes from microorganisms appear to be by far more suitable catalysts for bioconversion, although some different minor differences are present between them (e.g., a higher activity of the R. gracilis enzyme when the bioconversion is carried out at saturating oxygen concentration). The mammalian DAAO, even being a poor catalyst, is more stable with respect to temperature than the R. gracilis enzyme in the free form. In any case, for industrial purposes DAAO is used only in the immobilized form where a strong enzyme stabilization occurs.
Pollegioni
Loredano
L
Department of Structural and Functional Biology, University of Insubria, via J.H. Dunant 3, 21100 Varese, Italy.
Caldinelli
Laura
L
Molla
Gianluca
G
Sacchi
Silvia
S
Pilone
Mirella S
MS
D000818:Animals; D001711:Biotransformation; D002384:Catalysis; D002511:Cephalosporins / Q000096:biosynthesis* / Q000737:chemistry*; D003605:D-Amino-Acid Oxidase / Q000737:chemistry* / Q000145:classification*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000737:chemistry; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000201:enzymology*; D007700:Kinetics; D012248:Rhodotorula / Q000201:enzymology*; D004718:Saccharomycetales / Q000201:enzymology*; D013045:Species Specificity; D013379:Substrate Specificity; D013552:Swine; D013696:Temperature
D002511:Cephalosporins; D004791:Enzyme Inhibitors; C025163:cephalosporin C; C030735:7-aminocephalosporanic acid; D003605:D-Amino-Acid Oxidase