tetano
Editor, Senior Moderator
Virology
. 2021 Feb 9;557:15-22.
doi: 10.1016/j.virol.2021.01.004. Online ahead of print.
Expression, purification and immunological characterization of recombinant nucleocapsid protein fragment from SARS-CoV-2
Teodora Djukic[SUP] 1 [/SUP], Maja Mladenovic[SUP] 1 [/SUP], Dragana Stanic-Vucinic[SUP] 1 [/SUP], Jelena Radosavljevic[SUP] 1 [/SUP], Katarina Smiljanic[SUP] 1 [/SUP], Ljiljana Sabljic[SUP] 2 [/SUP], Marija Devic[SUP] 2 [/SUP], Danica Cujic[SUP] 2 [/SUP], Tamara Vasovic[SUP] 1 [/SUP], Ana Simovic[SUP] 1 [/SUP], Mirjana Radomirovic[SUP] 1 [/SUP], Tanja Cirkovic Velickovic[SUP] 3 [/SUP]
Affiliations
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.
Keywords: COVID-19; Prokaryotic expression; Recombinant nucleocapsid protein; SARS-CoV-2; serological assay.
. 2021 Feb 9;557:15-22.
doi: 10.1016/j.virol.2021.01.004. Online ahead of print.
Expression, purification and immunological characterization of recombinant nucleocapsid protein fragment from SARS-CoV-2
Teodora Djukic[SUP] 1 [/SUP], Maja Mladenovic[SUP] 1 [/SUP], Dragana Stanic-Vucinic[SUP] 1 [/SUP], Jelena Radosavljevic[SUP] 1 [/SUP], Katarina Smiljanic[SUP] 1 [/SUP], Ljiljana Sabljic[SUP] 2 [/SUP], Marija Devic[SUP] 2 [/SUP], Danica Cujic[SUP] 2 [/SUP], Tamara Vasovic[SUP] 1 [/SUP], Ana Simovic[SUP] 1 [/SUP], Mirjana Radomirovic[SUP] 1 [/SUP], Tanja Cirkovic Velickovic[SUP] 3 [/SUP]
Affiliations
- PMID: 33582454
- DOI: 10.1016/j.virol.2021.01.004
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.
Keywords: COVID-19; Prokaryotic expression; Recombinant nucleocapsid protein; SARS-CoV-2; serological assay.