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Improvement of catalytic efficiency and thermostability of recombinant Streptomyces griseus trypsin by introducing artificial peptide

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Abstract

Streptomyces trypsin is one of the serine proteinases in Streptomyces griseus and acts as a key mediator during cell growth and differentiation. S. griseus trypsin (SGT) could be successfully expressed in Pichia pastoris by engineering the natural propeptide APNP. In this study, the recombinant Exmt with peptide YVEF and the wild-type SGT were comparatively investigated in detail. The recombinant Exmt showed significantly increased thermostability which t 1/2 value was 3.89-fold of that of the SGT at 40 °C. Moreover, the catalytic efficiency (referring to the specificity constant, k cat/K m) and pH tolerance of Exmt were also improved. In silico modeling analysis uncovered that introduction of the peptide YVEF resulted in a broadened substrate binding pocket and closer catalytic triad (His57, Asp102 and Ser195). The intramolecular Hydrogen bonds and the cation π-interactions were also dramatically increased. The results indicated that engineering of the N-terminus with artificial peptides might be an effective approach for optimizing the properties of the target enzymes.

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Acknowledgments

This work was financially supported by the National High Technology Research and Development Program of China (863 Program, 2011AA100905), Program for Changjiang Scholars and Innovative Research Team in University (No. IRT1135), the National Natural Science Foundation of China (31200020, 31000031), the National Science Foundation for Post-doctoral Scientists of China (2013M540414) and the Jiangsu Planned Projects for Postdoctoral Research Funds (1301010B), the Doctor Candidate Foundation of Jiangnan University (JUDCF10014), the 111 Project and the Priority Academic Program Development of Jiangsu Higher Education Institutions.

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Correspondence to Zhen Kang or Guocheng Du.

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Ling, Z., Kang, Z., Liu, Y. et al. Improvement of catalytic efficiency and thermostability of recombinant Streptomyces griseus trypsin by introducing artificial peptide. World J Microbiol Biotechnol 30, 1819–1827 (2014). https://doi.org/10.1007/s11274-014-1608-1

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Keywords

  • Trypsin
  • Peptide
  • Catalytic efficiency
  • Thermostability