Abstract
Based on previous studies, two antibody-like molecules, monobodies, capable of high-affinity interaction with the SARS-CoV-2 nucleocapsid protein (dissociation constant of tens of nM) were selected. For delivery to target cells, genetically engineered constructs containing monobody and TAT peptide, placed either at the N- or C-terminus of the resulting polypeptide, were produced and expressed in E. coli. The construct with the highest affinity to the SARS-CoV-2 nucleocapsid protein was revealed with the use of thermophoresis technique. Cellular thermal shift assay demonstrated the ability of this construct to interact with the nucleocapsid protein within HEK293T cells transfected with the SARS-CoV-2 nucleocapsid protein fused to the mRuby3 fluorescent protein. Replacement of TAT peptide to S10 shuttle peptide, containing endosomolytic peptide, significantly improved the penetration of the construct into the target cells.
Keywords: S10 shuttle peptide; SARS-CoV-2; TAT peptide; antibody-like molecules; cellular thermal shift assay; monobody; nucleocapsid protein; thermophoresis.
【저자키워드】 SARS-CoV-2, nucleocapsid protein, antibody-like molecules, monobody, thermophoresis., cellular thermal shift assay, TAT peptide, S10 shuttle peptide, 【초록키워드】 peptide, Protein, nucleocapsid, cellular, Interaction, target cells, E. coli, Previous studies, fluorescent, penetration, C-terminus, replacement, highest affinity, TAT, selected, produced, resulting, significantly, expressed, demonstrated, fused, HEK293T cell, the SARS-CoV-2, transfected with, 【제목키워드】 delivery, target, molecules,