Expression, purification and immunological characterization of recombinant protein fragment from SARS-CoV-2
Аутори
Maja, MladenovićĐukić, Teodora
Vasović, Tamara
Stanić-Vučinić, Dragana
Smiljanić, Katarina
Radosavljević, Jelena
Sabljić, Ljiljana
Dević, Marija
Cujić, Danica
Ćirković-Veličković, Tanja
Конференцијски прилог (Објављена верзија)
Метаподаци
Приказ свих података о документуАпстракт
Serological testing is important method for diagnosis of severe acute respiratory syndrome coronavirus 2 (SARSCoV-2) infection. Nucleocapsid (N) protein is the most abundant virus derived protein and strong immunogen. We aimed to find its efficient, low-cost production, suitable for serological diagnosis. SARS-CoV-2 recombinant fragment of nucleocapsid protein (rfNP; 58–419 aa) was expressed in E. coli in soluble form, purified by immobilized metal ion affinity chromatography and strong cation exchange chromatography after which it was analyzed by Mass and CD spectrometry 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.
Кључне речи:
Recombinant nucleocapsid protein / COVID-19 / SARS-CoV-2 / Prokaryotic expression / serological assayИзвор:
FoodEnTwin Symposium: Novel analytical approaches in food and environmental sciences, June 16-18, 2021 Belgrade, Serbia, 2021, 33-33Финансирање / пројекти:
- 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".
- Министарство науке, технолошког развоја и иновација Републике Србије, институционално финансирање - 200168 (Универзитет у Београду, Хемијски факултет) (RS-MESTD-inst-2020-200168)
- FoodEnTwin-Twinning of research activities for the frontier research in the fields of food, nutrition and environmental omics (EU-H2020-810752)
- CAPSIDO – Developement of the assays for detection of SARS Cov-2 virus capsid proteins in biological fluids of COVID19 patients (RS-ScienceFundRS-Fond_2020_COVID19-7542203)
Напомена:
- Book of Abstracts
Колекције
Институција/група
Hemijski fakultet / Faculty of ChemistryTY - CONF AU - Maja, Mladenović AU - Đukić, Teodora AU - Vasović, Tamara AU - Stanić-Vučinić, Dragana AU - Smiljanić, Katarina AU - Radosavljević, Jelena AU - Sabljić, Ljiljana AU - Dević, Marija AU - Cujić, Danica AU - Ćirković-Veličković, Tanja PY - 2021 UR - http://cherry.chem.bg.ac.rs/handle/123456789/5737 AB - Serological testing is important method for diagnosis of severe acute respiratory syndrome coronavirus 2 (SARSCoV-2) infection. Nucleocapsid (N) protein is the most abundant virus derived protein and strong immunogen. We aimed to find its efficient, low-cost production, suitable for serological diagnosis. SARS-CoV-2 recombinant fragment of nucleocapsid protein (rfNP; 58–419 aa) was expressed in E. coli in soluble form, purified by immobilized metal ion affinity chromatography and strong cation exchange chromatography after which it was analyzed by Mass and CD spectrometry 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. C3 - FoodEnTwin Symposium: Novel analytical approaches in food and environmental sciences, June 16-18, 2021 Belgrade, Serbia T1 - Expression, purification and immunological characterization of recombinant protein fragment from SARS-CoV-2 SP - 33 EP - 33 UR - https://hdl.handle.net/21.15107/rcub_cherry_5737 ER -
@conference{ author = "Maja, Mladenović and Đukić, Teodora and Vasović, Tamara and Stanić-Vučinić, Dragana and Smiljanić, Katarina and Radosavljević, Jelena and Sabljić, Ljiljana and Dević, Marija and Cujić, Danica and Ćirković-Veličković, Tanja", year = "2021", abstract = "Serological testing is important method for diagnosis of severe acute respiratory syndrome coronavirus 2 (SARSCoV-2) infection. Nucleocapsid (N) protein is the most abundant virus derived protein and strong immunogen. We aimed to find its efficient, low-cost production, suitable for serological diagnosis. SARS-CoV-2 recombinant fragment of nucleocapsid protein (rfNP; 58–419 aa) was expressed in E. coli in soluble form, purified by immobilized metal ion affinity chromatography and strong cation exchange chromatography after which it was analyzed by Mass and CD spectrometry 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.", journal = "FoodEnTwin Symposium: Novel analytical approaches in food and environmental sciences, June 16-18, 2021 Belgrade, Serbia", title = "Expression, purification and immunological characterization of recombinant protein fragment from SARS-CoV-2", pages = "33-33", url = "https://hdl.handle.net/21.15107/rcub_cherry_5737" }
Maja, M., Đukić, T., Vasović, T., Stanić-Vučinić, D., Smiljanić, K., Radosavljević, J., Sabljić, L., Dević, M., Cujić, D.,& Ćirković-Veličković, T.. (2021). Expression, purification and immunological characterization of recombinant protein fragment from SARS-CoV-2. in FoodEnTwin Symposium: Novel analytical approaches in food and environmental sciences, June 16-18, 2021 Belgrade, Serbia, 33-33. https://hdl.handle.net/21.15107/rcub_cherry_5737
Maja M, Đukić T, Vasović T, Stanić-Vučinić D, Smiljanić K, Radosavljević J, Sabljić L, Dević M, Cujić D, Ćirković-Veličković T. Expression, purification and immunological characterization of recombinant protein fragment from SARS-CoV-2. in FoodEnTwin Symposium: Novel analytical approaches in food and environmental sciences, June 16-18, 2021 Belgrade, Serbia. 2021;:33-33. https://hdl.handle.net/21.15107/rcub_cherry_5737 .
Maja, Mladenović, Đukić, Teodora, Vasović, Tamara, Stanić-Vučinić, Dragana, Smiljanić, Katarina, Radosavljević, Jelena, Sabljić, Ljiljana, Dević, Marija, Cujić, Danica, Ćirković-Veličković, Tanja, "Expression, purification and immunological characterization of recombinant protein fragment from SARS-CoV-2" in FoodEnTwin Symposium: Novel analytical approaches in food and environmental sciences, June 16-18, 2021 Belgrade, Serbia (2021):33-33, https://hdl.handle.net/21.15107/rcub_cherry_5737 .