Generation of non-standard macrocyclic peptides specifically binding TSC-22 homologous gene-1.

Sophie T PhuongDung Tran, Christopher J Hipolito, Hiroyuki Suzuki, Rudy Xie, Huynh Dam Kim Tuyen, Peter Ten Dijke, Naohiro Terasaka, Yuki Goto, Hiroaki Suga, Mitsuyasu Kato

Journal: Biochemical and biophysical research communications 2020;516(2):445-450

PMID: 31227214

Full Text on Elsevier Science View on PubMed

Abstract

Transforming growth factor-β 1 (TGFβ1)-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.

Address: PhD Program in Human Biology, School of Integrative and Global Majors, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, 305-8575, Japan; Department of Experimental Pathology, Faculty of Medicine, University of Tsukuba, Japan.; Department of Cancer Signaling, Faculty of Medicine, University of Tsukuba, Japan; Transborder Medical Research Center, Faculty of Medicine, University of Tsukuba, Japan.; Department of Experimental Pathology, Faculty of Medicine, University of Tsukuba, Japan.; Department of Experimental Pathology, Faculty of Medicine, University of Tsukuba, Japan; Graduate School of Comprehensive Human Sciences, University of Tsukuba, Japan.; Department of Cancer Signaling, Faculty of Medicine, University of Tsukuba, Japan; Department of Molecular Cell and Chemical Biology, Oncode Institute, Leiden University Medical Center, Einthovenweg 20, 2333, ZC Leiden, the Netherlands.; Department of Chemistry, Graduate School of Science, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.; Department of Experimental Pathology, Faculty of Medicine, University of Tsukuba, Japan; Transborder Medical Research Center, Faculty of Medicine, University of Tsukuba, Japan. Electronic address: [email protected].

DOI: 10.1016/j.bbrc.2019.06.035

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MeSH Terms: Fluorescein, HEK293 Cells, Humans, Macrocyclic Compounds, Peptides, Transcription Factors