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Applied Microbiology and Biotechnology

, Volume 103, Issue 6, pp 2731–2743 | Cite as

Soluble Lactobacillus delbrueckii subsp. bulgaricus 92059 PrtB proteinase derivatives for production of bioactive peptide hydrolysates from casein

  • B. Li
  • D. Habermann
  • T. Kliche
  • M. Klempt
  • A. Wutkowski
  • I. Clawin-Rädecker
  • S. Koberg
  • E. Brinks
  • T. Koudelka
  • A. Tholey
  • W. BockelmannEmail author
  • C. M. A. P. Franz
  • K. J. Heller
Applied microbial and cell physiology

Abstract

The proteinase-encoding prtB gene of Lactobacillus (Lb.) delbrueckii (d.) subsp. bulgaricus 92059 was cloned and sequenced. Two soluble, secreted, C-terminally His-tagged derivatives were constructed and expressed in Lactococcus lactis by means of the NICE® Expression System. In both obtained derivatives PrtBb and PrtB2, the C-terminal, cell wall-binding domain was deleted. In addition, in derivative PrtB2, the C-terminal part of the B domain was deleted and the signal sequence was replaced by a lactococcal export signal. The affinity-purified derivatives were both proteolytically active. Peptide hydrolysates produced from casein with each of the derivatives showed identical peptide composition, as determined by liquid chromatography–mass spectrometry. Comparison of the peptides generated to those generated with living Lb. d. subsp. bulgaricus 92059 cells (Kliche et al. Appl Microbiol Biotechnol 101:7621–7633, 2017) showed that β-casein was the casein fraction most susceptible to hydrolysis and that some significant differences were observed between the products obtained by either the derivatives or living Lb. d. subsp. bulgaricus 92059 cells. When tested for biological activity, the hydrolysate obtained with PrtBb showed 50% inhibition of angiotensin-converting enzyme at a concentration of 0.5 mg/ml and immunomodulation/anti-inflammation in an in vitro assay of TNF-α induced NFκB activation at concentrations of 5 and 2.5 mg/ml, respectively. The enzymatically obtained hydrolysate did not show any pro-inflammatory or cytotoxic activity.

Keywords

Bioactive peptides Lactobacillus delbrueckii subsp. bulgaricus Proteinase Casein Antihypertensive activity Immunomodulatory activity 

Notes

Acknowledgements

Expert technical assistance by G. Gerke, R. Miller, F. Repenning and M. Steinke is gratefully acknowledged. T. Koudelka and A. Tholey were supported by Deutsche Forschungsgemeinschaft (DFG), within SFB877, project Z2.

Funding

The study was funded by the German Federal Ministry of Education and Research (Support Code 0315539B).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

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

Supplementary material

253_2018_9586_MOESM1_ESM.pdf (205 kb)
ESM 1 (PDF 205 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • B. Li
    • 1
  • D. Habermann
    • 1
  • T. Kliche
    • 1
  • M. Klempt
    • 2
  • A. Wutkowski
    • 2
  • I. Clawin-Rädecker
    • 2
  • S. Koberg
    • 1
  • E. Brinks
    • 1
  • T. Koudelka
    • 3
  • A. Tholey
    • 3
  • W. Bockelmann
    • 1
    Email author
  • C. M. A. P. Franz
    • 1
  • K. J. Heller
    • 1
  1. 1.Department of Microbiology and BiotechnologyMax Rubner-Institut (Federal Research Center of Nutrition and Food)KielGermany
  2. 2.Department of Safety and Quality of Milk and FishMax Rubner-Institut (Federal Research Center of Nutrition and Food)KielGermany
  3. 3.Systematic Proteomics and Bioanalytics, Institute of Experimental MedicineChristian-Albrechts-UniversityKielGermany

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