Immobilization of glutaryl-7-aminocephalosporanic acid acylase on silica gel and enhancement of its stability.
Glutaryl-7-aminocephalosporanic acid (GL-7-ACA) acylase is an enzyme that converts GL-7-ACA to 7-aminocephalosporanic acid, a starting material for semisynthetic cephalosporin antibiotics. In this study, optimal conditions for the immobilization of GL-7-ACA acylase were determined by experimental observations and statistical methods. The optimal conditions were as follows: 1.1 M phosphate buffer (pH 8.3) as buffer solution, immobilization temperature of 20 degrees C, and immobilization time of 120 min. Unreacted aldehyde groups were quenched by reaction with a low-molecular-weight material such as L-lysine, glycine, and ethanolamine after immobilization in order to enhance the activity of immobilized GL-7-ACA acylase. The activities of immobilized GL-7-ACA acylase obtained by using the low-molecular-weight materials were higher than those obtained by immobilized GL-7-ACA acylase not treated with low-molecular-weight materials. In particular, the highest activity of immobilized GL-7-ACA acylase was obtained using 0.4% (v/v) ethanolamine. We also investigated the effect of sodium cyanoborohydride in order to increase the stability of the linkage between the enzyme and the support. The effect on operational stability was obvious: the activity of immobilized GL-7-ACA acylase treated with 4% (w/w) sodium cyanoborohydride remained almost 100% after 20 times of reuse.
Park
Seung Won
SW
Department of Chemical and Biological Engineering, Korea University, 1 Anam-dong, Sungbuk-ku, Seoul 136-701, Korea.
Lee
Jee Won
JW
Hong
Suk In
SI
Kim
Seung Wook
SW
D001894:Borohydrides / Q000737:chemistry; D002511:Cephalosporins / Q000138:chemical synthesis / Q000737:chemistry; D004789:Enzyme Activation; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry*; D019856:Ethanolamine / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D008956:Models, Chemical; D010399:Penicillin Amidase / Q000737:chemistry*; D011786:Quality Control; D058428:Silica Gel; D012822:Silicon Dioxide / Q000737:chemistry*; D013696:Temperature
D001894:Borohydrides; D002511:Cephalosporins; D004800:Enzymes, Immobilized; C081063:glutaryl-7-aminocephalosporanic acid; D019856:Ethanolamine; D058428:Silica Gel; D012822:Silicon Dioxide; C030735:7-aminocephalosporanic acid; C009282:sodium cyanoborohydride; D010399:Penicillin Amidase; C046586:glutarylamidocephalosporanic acid acylase