Expression and secretion of SARS-CoV-2 antibody anti RBD single chain variable fragments in Saccharomyces cerevisiae

• Grant number: 23/03968-8
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: August 01, 2023
• End date: July 31, 2024
• Field of knowledge: Biological Sciences - Biochemistry - Molecular Biology
• Principal Investigator: Sandro Roberto Valentini
• Grantee: Yasmin Monteiro da Silva
• Host Institution: Faculdade de Ciências Farmacêuticas (FCFAR). Universidade Estadual Paulista (UNESP). Campus de Araraquara. Araraquara , SP, Brazil

Abstract

The COVID-19 pandemic has caused many impacts on society, emphasizing the hungry and the unemployment. Many restriction measures were taken with the purpose of reducing the virus dissemination speed and UTI beds overloads, such as social distancing and the mandatory use of masks. Inside this scenario, diagnostic kits sensitive to Sars-Cov-2 were broadly used as a way to track infected individuals, with the RT-qPCR being the standard test used to confirm the infection. However, this test demands a high quality technological structure, with proper equipment and a specialized workforce, besides being relatively expensive, which restricts its use. Furthermore, the lack of Brazilian autonomy in the production of the tests had become evident because of the external dependency on the active pharmaceutical supply, since there isn't a consolidated national production in this sector. That way, through this project it is intended to realize the expression and secretion of the antibody CC12.1 single-chain variable fragments (scFv), which has a high affinity to the Sars-Cov-2 receptor binding domain (RBD), in Saccharomyces cerevisiae, as well as its purification and examination of its functionality maintenance regarding the RBD. To accomplish that, the scFv coding sequence will be merged to the Aga2 glycoprotein, making the posterior cell secretion possible. After the transformation, induction and confirmation of the scFv expression in the yeast, the supernatant will be purified by the histidine tail present in the insert utilizing affinity chromatography and the fragments functionality maintenance will be evaluated by dot blot.