tANCHOR: A Novel Mammalian Cell Surface Peptide Display System

Abstract

A novel tool for the presentation of peptides and small proteins on the surface of human cells has been developed. Our tANCHOR system utilizes tetraspanin anchors containing heterologous amino acid sequences inserted instead of the large extracellular loop. This technology allows a highly effective extracellular display of epitopes for antibody binding studies and many other potential applications.

tANCHOR-cell-based assay for monitoring of SARS-CoV-2 neutralizing antibodies rapidly adaptive to various receptor-binding domains

“…We developed a cell-based ELISA for rapid and easy adaptation to upcoming SARS-CoV-2 variants by employing the tANCHOR protein display system. 21 This display system utilizes transmembrane anchors derived from Tspan sequences. Protein expression can be followed-up by fusion to the mCherry reporter protein ( Figure 1 A).…”

“…The hallmark of the tANCHOR system, especially for CD82-derived transmembrane domains, is its high efficiency for displaying proteins or peptides on the surface of human cells. 21 Therefore, we used this system to display the receptor-binding domain (RBD) of the Alpha (B.1.1.7), Beta (1.351), Gamma (P.1), Delta (B.1.617.2), Lambda (C.37), Omicron (BA.1), and the ancestral Wuhan (Wuhan-Hu-1) strain of SARS-CoV-2. Spike sequences corresponding to the amino acids (aa) 318–543 of the Wuhan-Hu-1 strain were defined as individual RBDs ( Figure 1 B).…”

“… 19 Importantly, the LEL is not necessary for routing a Tspan to the cell surface and can be replaced by heterologous protein sequences. 20 , 21 In particular, the transmembrane anchors derived from the Tspan CD82 showed best performance for displaying proteins on the cell surface of human embryonic kidney 293T (HEK293T) or HeLa cells, where the protein of interest is fused and expressed as a chimeric-membrane-bound unit connected by optimized linker sequences between the third and fourth TM. 21 , 22 For testing the neutralization activity of an SARS-CoV-2 variant, only the coding DNA sequence in the tANCHOR vectors has to be adjusted.…”

Establishment of a human cell line with a surface display system for screening and optimizing Na+-taurocholate cotransporting polypeptide-binding peptides

“…To tackle these challenges, surface display strategies based on human cells have been developed recently ( Crook et al, 2017 ; Chan et al, 2020 ; Ivanusic et al, 2020 ). Here, we set up a system suitable for the screening of NTCP-binding peptides.…”

Quantum Dots Tracking Endocytosis and Transport of Proteins Displayed by Mammalian Cells

“…Surface display technology uses recombinant DNA technology to express and localize exogenous peptides or proteins on the cell surface. − In contrast to phage display technology and bacterial surface display technology, mammalian cell display technology can guide protein folding and provide various post-translational processing functions, resulting in expressed products that are closest to natural biological protein molecules in terms of molecular structure, physicochemical properties, and biological function. , This technology has been successfully applied to directed evolution of enzymes, peptide library screening, high-affinity antibody sorting, and antigen/antibody library construction, , and new applications continue to emerge. Current studies on mammalian cell display technology are focused on the function of the displayed protein, with less emphasis on the “fate” of these proteins after they are displayed on the cell membrane.…”