Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/42103
標題: Simultaneous purification and immobilization of d-hydantoinase on the immobilized metal affinity membrane via coordination bonds
作者: Ko, Yi-Miao
Chen, Chih-I
Shieh, Chwen-Jen
Liu, Yung-Chuan
關鍵字: d-Hydantoinase;Immobilized metal affinity membrane;Enzyme immobilization;Enzyme purification;Enzyme stability
Project: Biochemical Engineering Journal, Volume 61, page(s) 20– 27.
摘要: 
This study constructs the immobilized metal affinity membrane (IMAM) via coupling of epichlorohydrin,
iminodiacetic acid, and nickel ion on the regenerated cellulose membrane. The d-hydantoin-hydrolyzing
enzyme (DHTase) harboring a poly-His tagged residue was used as a model protein immobilized on the
prepared IMAM. Various immobilization conditions were examined based on the yield of N-carbamoyl-dp-
hydroxyphenylglycine in batch reactions. The immobilization conditions were studied and the optimal
conditions are as follows. By employing an IMAM with nickel ion of 155.5 ± 5 !mol/disc immersed in 0.1 M
Tris–HCl buffer pH 8 (with 0.8 M sodium chloride) and immobilized time of 14 h, a DHTase activity of
4.2 ± 0.3 U/disc was obtained. The immobilized DHTase membrane can achieve a larger pH and thermal
tolerant range than that of free enzyme. Meanwhile, the stability test showed that 99% of enzyme activity
could be retained after being repeated 15-times. The storage test also displayed 99% enzyme preservation
after 7 weeks of storage.
URI: http://hdl.handle.net/11455/42103
ISSN: 1369-703X
DOI: 10.1016/j.bej.2011.11.013
Appears in Collections:化學工程學系所

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