Affinity induced immobilization of adenylate cyclase from the crude cell lysate for ATP conversion

Qianqian Zhu, Wei Zhuang, Huanqing Niu, Lei Ge, Byron Villacorta Hernandez, Jinglan Wu, Kai Wang, Dong Liu, Yong Chen, Chenjie Zhu, Hanjie Ying

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

The development of an orientation immobilization technique via affinity between polyhistidine tags and metal ions aims at maintaining biocatalytic activity of the enzymes. In this work, to tackle the issue of the immobilization of adenylate cyclase (AC), a simple and effective approach of synthesizing iminodiacetic acid (IDA)-Ni2+ particles was applied for simultaneously purifying and immobilizing his-tagged AC. We chose agarose particles as carriers, and then decorated them with IDA, leading to the formation of a coordination combination of Ni2+. The porous carriers with a large pore size of 50 nm and a specific surface area of 45.8 m2/g exhibited favorable enzymatic activity and loading capacity. The optimal pH of the immobilized enzyme increased from 8.0 to 9.0 and the optimal temperature increased from 30 °C to 35 °C, compared to the free AC. Moreover, the immobilized AC retained a residual activity of approximately 80% after storing it at 25 °C for 48 h, whereas only 40% of the activity was left in the free AC at the same conditions. Maximum yield of cyclic adenosine-3′, 5′- monophosphate (cAMP) reached up to the summit of the reaction. The immobilized AC by affinity adsorption will provide a promising route for the industrial production of cAMP.

Original languageEnglish
Pages (from-to)155-164
Number of pages10
JournalColloids and Surfaces B: Biointerfaces
Volume164
DOIs
StatePublished - 1 Apr 2018

Keywords

  • Adenylate cyclase
  • Cyclic adenosine-3′,5′-monophosphate
  • Enzyme immobilization
  • His-tagged
  • Iminodiacetic acid
  • Nickel ions

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