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High Production of Neuraminidase by a Vibrio cholerae Non-O1 Strain—the First Possible Alternative to Toxigenic Producers

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Abstract

Vibrio cholerae neuraminidase (VCNA) is widely used in biochemical and medical research, in processes for preparing homogenous sialoconjugates, and in the pharmaceutical industry. Its production by non-toxigenic strains is quite desirable, in order to avoid the expensive safety measures. Here, we report the first method for highly effective production of a novel, purified V. cholerae extracellular neuraminidase from a non-toxigenic strain. The enzyme is highly active, and its properties, as well as the responsible gene nanH, are practically identical with those of the toxigenic strains. It cleaves α,2 → 3 and α,2 → 6 glycosidic bonds with highest affinity (K M 1.7 × 10−5 μM) for human transferrin. The deduced amino acid sequence of the enzyme reveals three binding sites for Ca2+ and one for sialic acid. The sequence analysis of the nanH gene, being the first for a V. cholerae non-O1 strain, shows 99 % identity with a new nanH allele of an O1 Vibrio strain. The simple laboratory technology for efficient production of the new VCNA is based on the use of common and cheap nutrient media and easily available inducer—glycomacropeptide. The rapid purification consists of salting-out and diethylaminoethanol (DEAE) and Q-Sepharose chromatography steps. Purified preparation contains no aldolase and protease, which gives the production scheme a great potential for industrial application.

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Acknowledgments

This work was supported by grant G-2-02/05 from the Ministry of Education and Science, Science Research Fund, Bulgaria. We thank Mr. Dejan Draganov for providing 4MU-Neu5Ac substrate and research assistant Anna Bratchkova for the visualization of the native PAGE.

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Authors and Affiliations

  1. The Stephan Angeloff Istitute of Microbiology, Sofia, Bulgaria

    Rumyana Todorova Eneva, Stephan Andreas Engibarov, Penka Petrova, Radoslav Abrashev & Ignat Abrashev

  2. Department of Microbiology, Medical University, Sofia, Bulgaria

    Tanya Strateva

  3. Institute of Experimental Morphology, Pathology and Anthropology with Museum, Bulgarian Academy of Sciences, Sofia, Bulgaria

    Vera Kolyovska

Authors
  1. Rumyana Todorova Eneva
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  2. Stephan Andreas Engibarov
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  3. Penka Petrova
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  4. Radoslav Abrashev
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  5. Tanya Strateva
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  6. Vera Kolyovska
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  7. Ignat Abrashev
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Corresponding author

Correspondence to Rumyana Todorova Eneva.

Additional information

Stephan Andreas Engibarov holds a MS degree.

Penka Petrova holds a PhD degree and is an Associated professor of The Stephan Angeloff Institute of Microbiology.

Radoslav Abrashev holds a PhD degree.

Tanya Strateva, PhD, is the Associated professor of The Department of Microbiology, Medical University, Sofia.

Vera Kolyovska holds a PhD degree.

Ignat Abrashev, PhD, is the Professor, DSc of The Stephan Angeloff Istitute of Microbiology.

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Eneva, R.T., Engibarov, S.A., Petrova, P. et al. High Production of Neuraminidase by a Vibrio cholerae Non-O1 Strain—the First Possible Alternative to Toxigenic Producers. Appl Biochem Biotechnol 176, 412–427 (2015). https://doi.org/10.1007/s12010-015-1584-4

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  • DOI: https://doi.org/10.1007/s12010-015-1584-4

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