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Synthesis of human parainfluenza virus 4 nucleocapsid-like particles in yeast and their use for detection of virus-specific antibodies in human serum

  • Applied genetics and molecular biotechnology
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Abstract

The aim of this study was to produce human parainfluenza virus type 4 (HPIV4) nucleocapsid (N) protein in yeast Saccharomyces cerevisiae expression system, to explore its structural and antigenic properties and to evaluate its applicability in serology. The use of an optimized gene encoding HPIV4 N protein amino acid (aa) sequence GenBank AGU90031.1 allowed high yield of recombinant N protein forming nucleocapsid-like particles (NLPs) in yeast. A substitution L332D disrupted self-assembly of NLPs, confirming the role of this position in the N proteins of Paramyxovirinae. Three monoclonal antibodies (MAbs) were generated against the NLP-forming HPIV4 N protein. They recognised HPIV4-infected cells, demonstrating the antigenic similarity between the recombinant and virus-derived N proteins. HPIV4 N protein was used as a coating antigen in an indirect IgG ELISA with serum specimens of 154 patients with respiratory tract infection. The same serum specimens were tested with previously generated N protein of a closely related HPIV2, another representative of genus Rubulavirus. Competitive ELISA was developed using related yeast-produced viral antigens to deplete the cross-reactive serum antibodies. In the ELISA either without or with competition using heterologous HPIV (2 or 4) N or mumps virus N proteins, the seroprevalence of HPIV4 N-specific IgG was, respectively, 46.8, 39.6 and 40.3% and the seroprevalence of HPIV2 N-specific IgG—47.4, 39.0 and 37.7%. In conclusion, yeast-produced HPIV4 N protein shares structural and antigenic properties of the native virus nucleocapsids. Yeast-produced HPIV4 and HPIV2 NLPs are prospective tools in serology.

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Acknowledgements

We thank Dr. Regina Ėmužytė and Dr. Regina Firantienė, who provided clinical data and patient serum specimens collected at the Clinic of Children’s Diseases, Faculty of Medicine, Vilnius University (Lithuania).

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  1. Institute of Biotechnology, Vilnius University, Saulėtekio 7, LT-10257, Vilnius, Lithuania

    Aistė Bulavaitė, Rita Lasickienė, Paulius Lukas Tamošiūnas, Martynas Simanavičius, Kęstutis Sasnauskas & Aurelija Žvirblienė

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  1. Aistė Bulavaitė
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  2. Rita Lasickienė
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  3. Paulius Lukas Tamošiūnas
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Correspondence to Aistė Bulavaitė.

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The current study did not receive any specific grant from a funding agency in the public, commercial or not for-profit sectors.

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The authors declare that they have no conflict of interest.

Animal welfare

All applicable institutional, national and international guidelines for the care and use of animals were followed. The mice were used under the licence No. LT 59-902, permission No. 184 for breeding of experimental mice and permission No. 209 for generation of polyclonal and monoclonal antibodies.

Human participants

Collection of patients’ clinical data and serum specimens was performed with informed consent following applicable ethical guidelines and was approved by the Lithuanian Bioethics Committee (No. 53, 2008-09-03).

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Bulavaitė, A., Lasickienė, R., Tamošiūnas, P.L. et al. Synthesis of human parainfluenza virus 4 nucleocapsid-like particles in yeast and their use for detection of virus-specific antibodies in human serum. Appl Microbiol Biotechnol 101, 2991–3004 (2017). https://doi.org/10.1007/s00253-017-8104-0

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