o:25895 Expression, purification and immunological characterization of recombinant nucleocapsid protein fragment from SARS-CoV-2 Ekspresija, prečišćavanje i imunološka karakterizacija fragmenta rekombinantnog proteina nukleokapsida SARS-COV-2 en 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. recombinant nucleocapsid protein, COVID-19, SARS-CoV-2, prokaryotic expression, serological assay 1552099 10.1016/j.virol.2021.01.004 2022-06-03T10:38:22.718Z 44 yes 46 Teodora Djukić University of Belgrade, Faculty of Chemistry, Department of Biochemistry, Centre of Excellence for Molecular Food Sciences, Serbia Tanja Ćirković Veličković University of Belgrade, Faculty of Chemistry, Department of Biochemistry, Centre of Excellence for Molecular Food Sciences, Serbia person Mirjana Radomirović University of Belgrade, Faculty of Chemistry, Department of Biochemistry, Centre of Excellence for Molecular Food Sciences, Serbia person Ana Simović University of Belgrade, Faculty of Chemistry, Department of Biochemistry, Centre of Excellence for Molecular Food Sciences, Serbia person Tamara Vasović University of Belgrade, Faculty of Chemistry, Department of Biochemistry, Centre of Excellence for Molecular Food Sciences, Serbia person Danica Ćujić University of Belgrade, Institute for the Application of Nuclear Energy INEP, Serbia doktor medicinskih nauka-farmacija person 0000-0002-6145-3850 Marija Dević University of Belgrade, Institute for the Application of Nuclear Energy INEP, Serbia doktor biotehničkih nauka person 0000-0001-8233-1636 Ljiljana Sabljić University of Belgrade, Institute for the Application of Nuclear Energy INEP, Serbia person 0000-0001-6027-591X Katarina Smiljanić University of Belgrade, Faculty of Chemistry, Department of Biochemistry, Centre of Excellence for Molecular Food Sciences, Serbia person Jelena Radosavljević University of Belgrade, Faculty of Chemistry, Department of Biochemistry, Centre of Excellence for Molecular Food Sciences, Serbia person Dragana Stanić-Vučinić University of Belgrade, Faculty of Chemistry, Department of Biochemistry, Centre of Excellence for Molecular Food Sciences, Serbia person Maja Mladenović University of Belgrade, Faculty of Chemistry, Department of Biochemistry, Centre of Excellence for Molecular Food Sciences, Serbia person application/pdf 3913496 https://phaidrabg.bg.ac.rs/o:25895 no yes 1 70 92000001 11A19 11A1903 11A29 11A2904 Virology 557 15 22 Elsevier 2021