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Molecular Modeling and Expression of Multimeric Bann-RBD Fusion Protein in Pichia pastoris as a COVID-19 Vaccine Candidate (a) Institut Teknologi Bandung, Indonesia Abstract The COVID-19 pandemic caused by the transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has a wide impact on health, economy, and other sectors. Various vaccine platforms have been developed and are being administered worldwide to provide protection. Subunit protein vaccines are one of the most widely developed COVID-19 vaccine candidates. The receptor binding domain (RBD) of spike protein has been used as an antigen as it can stimulate the immune system to produce neutralizing antibodies that can inhibit the interaction of the virus with ACE2. Several studies have shown that dimeric, trimeric, and multimeric RBD produce higher neutralizing antibodies than monomeric RBD. In this study, we have developed a multimeric RBD as a protein fusion of RBD and β--annulus (Bann) from tomato bushy stunt virus (TBSV). Pichia pastoris X-33 was chosen as an expression host because it can secrete protein efficiently and has a post-translational modification system. The molecular modeling of the Bann-RBD protein shows a 60-mer structure with Bann being at the core and RBD being displayed on the surface. P. pastoris X-33-pPICZα-A-Bann-RBD has been successfully constructed. SDS PAGE analysis of the Bann-RBD protein expressed in P. pastoris X-33 showed several protein bands which are predicted to be monomer, and multimer of glycosylated Bann-RBD. Further characterization indicated that the resulted recombinant Bann-RBD can recognize strongly an antibody of COVID-19 in positive patient serum. Taken together, this results indicate that the recombinant Bann-RBD produced in P. pastoris X-33 could be used in the development of COVID-19 candidate vaccine. Keywords: Keywords: COVID-19- multimeric protein- RBD- subunit vaccine- β--annulus Topic: Biokimia Medis |
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