A Soluble, Minimalistic Glycosylphosphatidylinositol Transamidase (GPI-T) Retains Transamidation Activity.

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  • Additional Information
    • Source:
      Publisher: American Chemical Society Country of Publication: United States NLM ID: 0370623 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1520-4995 (Electronic) Linking ISSN: 00062960 NLM ISO Abbreviation: Biochemistry Subsets: MEDLINE
    • Publication Information:
      Original Publication: Washington, American Chemical Society.
    • Subject Terms:
      Acyltransferases*/chemistry ; Acyltransferases*/metabolism ; Saccharomyces cerevisiae Proteins*/metabolism; Glycosylphosphatidylinositols ; Humans ; Saccharomyces cerevisiae/enzymology
    • Abstract:
      Glycosylphosphatidylinositol (GPI) anchoring of proteins is a eukaryotic, post-translational modification catalyzed by GPI transamidase (GPI-T). The Saccharomyces cerevisiae GPI-T is composed of five membrane-bound subunits: Gpi8, Gaa1, Gpi16, Gpi17, and Gab1. GPI-T has been recalcitrant to in vitro structure and function studies because of its complexity and membrane-solubility. Furthermore, a reliable, quantitative, in vitro assay for this important post-translational modification has remained elusive despite its discovery more than three decades ago.Three recent reports describe the structure of GPI-T from S. cerevisiae and humans, shedding critical light on this important enzyme and offering insight into the functions of its different subunits. Here, we present the purification and characterization of a truncated soluble GPI-T heterotrimer complex (Gpi8 23-306 , Gaa1 50-343 , and Gpi16 20-551 ) without transmembrane domains. Using this simplified heterotrimer, we report the first quantitative method to measure GPI-T activity in vitro and demonstrate that this soluble, minimalistic GPI-T retains transamidase activity. These results contribute to our understanding of how this enzyme is organized and functions, and provide a method to screen potential GPI-T inhibitors.
    • Accession Number:
      0 (Glycosylphosphatidylinositols)
      0 (Saccharomyces cerevisiae Proteins)
      EC 2.3.- (Acyltransferases)
      EC 2.3.2.- (COOH-terminal signal transamidase)
    • Publication Date:
      Date Created: 20220622 Date Completed: 20220707 Latest Revision: 20220725
    • Publication Date:
      20231215
    • Accession Number:
      10.1021/acs.biochem.2c00196
    • Accession Number:
      35730892