Expression, purification and immunological characterization of recombinant nucleocapsid protein fragment from SARS-CoV-2

Serological testing is important method for diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Nucleocapsid (N) protein is the most abundant virus derived protein and strong immunogen. We aimed to find its efficient, low-cost production. SARS-CoV-2 recombinant fragment of nucleocapsid protein (rfNP; 58–419 aa) was expressed in E. coli in soluble form, purified and characterized biochemically and immunologically. Purified rfNP has secondary structure of full-length recombinant N protein, with high percentage of disordered structure (34.2%) and of β-sheet (40.7%). rfNP was tested in immunoblot using sera of COVID-19 convalescent patients. ELISA was optimized with sera of RT-PCR confirmed positive symptomatic patients and healthy individuals. IgG detection sensitivity was 96% (47/50) and specificity 97% (67/68), while IgM detection was slightly lower (94% and 96.5%, respectively). Cost-effective approach for soluble recombinant N protein fragment...

Keywords:

Recombinant nucleocapsid protein / COVID-19 / SARS-CoV-2 / Prokaryotic expression / serological assay

Source:
Virology journal, 2021, 557, 15-22

Publisher:

Elsevier

Funding / projects:

CAPSIDO – Developement of the assays for detection of SARS Cov-2 virus capsid proteins in biological fluids of COVID19 patients (RS-7542203)
Ministry of Education, Science and Technological Development, Republic of Serbia, Grant no. 200168 (University of Belgrade, Faculty of Chemistry) (RS-200168)
Serbian Academy of Sciences and Arts GA No. F-26.
UNDP project "Preventing and Responding to COVID-19 in At-risk areas - Sustainable production of the serological IgG test for SARS CoV-2 in Serbia".

Note:

This is the preprint of the article: Đukić, T.; Mladenović, M.; Stanić-Vučinić, D.; Radosavljević, J.; Smiljanić, K.; Sabljić, L.; Dević, M.; Ćujić, D.; Vasović, T.; Simović, A.; Radomirović, M. Ž.; Ćirković-Veličković, T. Expression, Purification and Immunological Characterization of Recombinant Nucleocapsid Protein Fragment from SARS-CoV-2. Virology journal 2021, 557, 15–22. https://doi.org/10.1016/j.virol.2021.01.004.

Related info:

Version of
https://cherry.chem.bg.ac.rs/handle/123456789/4331

DOI: 10.1016/j.virol.2021.01.004

ISSN: 0042-6822

WoS: 000632032400001

Scopus: 2-s2.0-85100810713

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URI

https://cherry.chem.bg.ac.rs/handle/123456789/4330

Collections

Institution/Community

Hemijski fakultet / Faculty of Chemistry

TY  - JOUR
AU  - Đukić, Teodora
AU  - Mladenović, Maja
AU  - Stanić-Vučinić, Dragana
AU  - Radosavljević, Jelena
AU  - Smiljanić, Katarina
AU  - Sabljić, Ljiljana
AU  - Dević, Marija
AU  - Ćujić, Danica R.
AU  - Vasović, Tamara
AU  - Simović, Ana
AU  - Radomirović, Mirjana Ž.
AU  - Ćirković-Veličković, Tanja
PY  - 2021
UR  - https://cherry.chem.bg.ac.rs/handle/123456789/4330
AB  - Serological testing is important method for diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Nucleocapsid (N) protein is the most abundant virus derived protein and strong immunogen. We aimed to find its efficient, low-cost production.

SARS-CoV-2 recombinant fragment of nucleocapsid protein (rfNP; 58–419 aa) was expressed in E. coli in soluble form, purified and characterized biochemically and immunologically.

Purified rfNP has secondary structure of full-length recombinant N protein, with high percentage of disordered structure (34.2%) and of β-sheet (40.7%). rfNP was tested in immunoblot using sera of COVID-19 convalescent patients. ELISA was optimized with sera of RT-PCR confirmed positive symptomatic patients and healthy individuals. IgG detection sensitivity was 96% (47/50) and specificity 97% (67/68), while IgM detection was slightly lower (94% and 96.5%, respectively).

Cost-effective approach for soluble recombinant N protein fragment production was developed, with reliable IgG and IgM antibodies detection of SARS-CoV-2 infection.
PB  - Elsevier
T2  - Virology journal
T1  - Expression, purification and immunological characterization of recombinant nucleocapsid protein fragment from SARS-CoV-2
VL  - 557
SP  - 15
EP  - 22
DO  - 10.1016/j.virol.2021.01.004
ER  -

@article{
author = "Đukić, Teodora and Mladenović, Maja and Stanić-Vučinić, Dragana and Radosavljević, Jelena and Smiljanić, Katarina and Sabljić, Ljiljana and Dević, Marija and Ćujić, Danica R. and Vasović, Tamara and Simović, Ana and Radomirović, Mirjana Ž. and Ćirković-Veličković, Tanja",
year = "2021",
abstract = "Serological testing is important method for diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Nucleocapsid (N) protein is the most abundant virus derived protein and strong immunogen. We aimed to find its efficient, low-cost production.

SARS-CoV-2 recombinant fragment of nucleocapsid protein (rfNP; 58–419 aa) was expressed in E. coli in soluble form, purified and characterized biochemically and immunologically.

Purified rfNP has secondary structure of full-length recombinant N protein, with high percentage of disordered structure (34.2%) and of β-sheet (40.7%). rfNP was tested in immunoblot using sera of COVID-19 convalescent patients. ELISA was optimized with sera of RT-PCR confirmed positive symptomatic patients and healthy individuals. IgG detection sensitivity was 96% (47/50) and specificity 97% (67/68), while IgM detection was slightly lower (94% and 96.5%, respectively).

Cost-effective approach for soluble recombinant N protein fragment production was developed, with reliable IgG and IgM antibodies detection of SARS-CoV-2 infection.",
publisher = "Elsevier",
journal = "Virology journal",
title = "Expression, purification and immunological characterization of recombinant nucleocapsid protein fragment from SARS-CoV-2",
volume = "557",
pages = "15-22",
doi = "10.1016/j.virol.2021.01.004"
}

Đukić, T., Mladenović, M., Stanić-Vučinić, D., Radosavljević, J., Smiljanić, K., Sabljić, L., Dević, M., Ćujić, D. R., Vasović, T., Simović, A., Radomirović, M. Ž.,& Ćirković-Veličković, T.. (2021). Expression, purification and immunological characterization of recombinant nucleocapsid protein fragment from SARS-CoV-2. in Virology journal
Elsevier., 557, 15-22.
https://doi.org/10.1016/j.virol.2021.01.004

Đukić T, Mladenović M, Stanić-Vučinić D, Radosavljević J, Smiljanić K, Sabljić L, Dević M, Ćujić DR, Vasović T, Simović A, Radomirović MŽ, Ćirković-Veličković T. Expression, purification and immunological characterization of recombinant nucleocapsid protein fragment from SARS-CoV-2. in Virology journal. 2021;557:15-22.
doi:10.1016/j.virol.2021.01.004 .

Đukić, Teodora, Mladenović, Maja, Stanić-Vučinić, Dragana, Radosavljević, Jelena, Smiljanić, Katarina, Sabljić, Ljiljana, Dević, Marija, Ćujić, Danica R., Vasović, Tamara, Simović, Ana, Radomirović, Mirjana Ž., Ćirković-Veličković, Tanja, "Expression, purification and immunological characterization of recombinant nucleocapsid protein fragment from SARS-CoV-2" in Virology journal, 557 (2021):15-22,
https://doi.org/10.1016/j.virol.2021.01.004 . .