Oriented Immobilization and Characterization of a Poly-Lysine-Tagged Cephalosporin C Acylase on Glyoxyl Agarose Support

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• 作者：Hui Luo ; Huan Zhao ; Yanhong Chang ; Qixin Wang…
• 关键词：Cephalosporin C acylase ; Characterization ; Enzyme immobilization ; Genetic modification ; Glyoxyl agarose ; Poly ; lysine tag
• 刊名：Applied Biochemistry and Biotechnology
• 出版年：2015
• 出版时间：February 2015
• 年：2015
• 卷：175
• 期：4
• 页码：2114-2123
• 全文大小：503 KB
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• 刊物类别：Chemistry and Materials Science
• 刊物主题：Chemistry
  Biotechnology
  Biochemistry
  
• 出版者：Humana Press Inc.
• ISSN：1559-0291

文摘

Cephalosporin C acylase (CCA), an important industrial enzyme for the production of 7-aminocephalosporanic acid, has very limited and scattered surface lysine residues. A mutant of cephalosporin C acylase (mCCA) has been designed to fuse a poly-lysine tag to the C-terminal of the β-subunit, which is far away from the active site. The free mCCA showed a near equal specific activity with the wild-type CCA, while a much higher activity recovery was obtained for the mCCA than its wild-type counterpart after immobilization on glyoxyl agarose support (73.3 versus 53.3?%). The mCCA’s oriented immobilization enables it to obtain a higher substrate affinity and even a higher thermal stability than the wild-type enzyme. The improvement of stability might be attributed to the multipoint covalent attachment by the oriented enzyme immobilization via the adhered poly-lysine tag, which prevents the dissociation of the β-subunit of CCA from the support.