Applied Microbiology and Biotechnology

, Volume 100, Issue 18, pp 8063–8074 | Cite as

Enhanced exo-inulinase activity and stability by fusion of an inulin-binding module

  • Shun-Hua Zhou
  • Yuan Liu
  • Yu-Juan Zhao
  • Zhe Chi
  • Zhen-Ming Chi
  • Guang-Lei LiuEmail author
Biotechnologically relevant enzymes and proteins


In this study, an inulin-binding module from Bacillus macerans was successfully fused to an exo-inulinase from Kluyveromyces marxianus, creating a hybrid functional enzyme. The recombinant exo-inulinase (rINU), the hybrid enzyme (rINUIBM), and the recombinant inulin-binding module (rIBM) were, respectively, heterologously expressed and biochemically characterized. It was found that both the inulinase activity and the catalytic efficiency (k cat/K m(app)) of the rINUIBM were considerably higher than those of rINU. Though the rINU and the rINUIBM shared the same optimum pH of 4.5, the optimum temperature of the rINUIBM (60 °C) was 5 °C higher than that of the rINU. Notably, the fused IBM significantly enhanced both the pH stability and the thermostability of the rINUIBM, suggesting that the rINUIBM obtained would have more extensive potential applications. Furthermore, the fusion of the IBM could substantially improve the inulin-binding capability of the rINUIBM, which was consistent with the determination of the K m(app). This meant that the fused IBM could play a critical role in the recognition of polysaccharides and enhanced the hydrolase activity of the associated inulinase by increasing enzyme-substrate proximity. Besides, the extra supplement of the independent non-catalytic rIBM could also improve the inulinase activity of the rINU. However, this improvement was much better in case of the fusion. Consequently, the IBM could be designated as a multifunctional domain that was responsible for the activity enhancement, the stabilization, and the substrate binding of the rINUIBM. All these features obtained in this study make the rINUIBM become an attractive candidate for an efficient inulin hydrolysis.


Inulin-binding module Exo-inulinase pH stability Thermostability Inulin-binding capability Protein engineering 



This research was supported by the National Natural Science Foundation of China (Grant no. 31400047) and Key Research and Development Plan of Shandong Province (Grant no. 2015GSF121024).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

253_2016_7587_MOESM1_ESM.pdf (750 kb)
ESM 1 (PDF 749 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Shun-Hua Zhou
    • 1
  • Yuan Liu
    • 1
  • Yu-Juan Zhao
    • 1
  • Zhe Chi
    • 1
  • Zhen-Ming Chi
    • 1
  • Guang-Lei Liu
    • 1
    Email author
  1. 1.College of Marine Life ScienceOcean University of ChinaQingdaoChina

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