RESUMO
Transforming growth factor-ß 1 (TGFß1)-stimulated clone 22 (TSC22) family includes proteins containing a leucine zipper domain and a TSC-box that are highly conserved during evolution. Currently, limited data are available on the function of this protein family, especially of TSC-22 homologous gene-1 (THG-1)/TSC22 domain family member 4 (TSC22D4). Similar to other family members, THG-1 functions depending on its interaction with the partner proteins and it is suggested to mediate a broad range of biological processes. THG-1-specific binding molecules will be instrumental for elucidating its functions. Therefore, the Random non-standard Peptide Integrated Discovery (RaPID) system was modified using commercially available materials and used for selecting macrocyclic peptides (MCPs) that bind to THG-1. Several MCPs were identified to bind THG-1. Fluorescein- and biotin-tagged MCPs were synthesized and employed as THG-1 detection probes. Notably, a fluorescein-tagged MCP specifically detected THG-1-expressing cells. Biotin-tagged MCPs can be successfully used for Enzyme-Linked Protein Sorbent Assay (ELISA) like assay of THG-1 protein and affinity-precipitation of purified THG-1 and endogenous THG-1 in esophageal squamous cell carcinoma cell lysates. The modified RaPID system rapidly and successfully identified THG-1-binding MCPs in vitro and the synthesized THG-1 binding MCPs are useful alternatives acting for antibodies.
