Delving into human α1,4-galactosyltransferase acceptor specificity: The role of enzyme dimerization.

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  • Additional Information
    • Source:
      Publisher: Elsevier Country of Publication: United States NLM ID: 0372516 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1090-2104 (Electronic) Linking ISSN: 0006291X NLM ISO Abbreviation: Biochem Biophys Res Commun Subsets: MEDLINE
    • Publication Information:
      Publication: <2002- >: San Diego, CA : Elsevier
      Original Publication: New York, Academic Press.
    • Subject Terms:
      Galactosyltransferases*/metabolism ; Galactosyltransferases*/chemistry ; Galactosyltransferases*/genetics ; Cricetulus* ; Protein Multimerization*; Humans ; Animals ; CHO Cells ; HEK293 Cells ; Substrate Specificity ; Globosides/metabolism ; Globosides/chemistry ; Glycosphingolipids/metabolism ; Glycosphingolipids/chemistry
    • Abstract:
      Human α1,4-galactosyltransferase (A4galt), a Golgi apparatus-resident GT, synthesizes Gb3 glycosphingolipid (GSL) and P1 glycotope on glycoproteins (GPs), which are receptors for Shiga toxin types 1 and 2. Despite the significant role of A4galt in glycosylation processes, the molecular mechanisms underlying its varied acceptor specificities remain poorly understood. Here, we attempted to elucidate A4galt specificity towards GSLs and GPs by exploring its interaction with GTs with various acceptor specificities, GP-specific β1,4-galactosyltransferase 1 (B4galt1) and GSL-specific β1,4-galactosyltransferase isoenzymes 5 and 6 (B4galt5 and B4galt6). Using a novel NanoBiT assay, we found that A4galt can form homodimers and heterodimers with B4galt1 and B4galt5 in two cell lines, human embryonic kidney cells (HEK293T) and Chinese hamster ovary cells (CHO-Lec2). We found that A4galt-B4galts heterodimers preferred N-terminally tagged interactions, while in A4galt homodimers, the favored localization of the fused tag depended on the cell line used. Furthermore, by employing AlphaFold for state-of-the-art structural prediction, we analyzed the interactions and structures of these enzyme complexes. Our analysis highlighted that the A4galt-B4galt5 heterodimer exhibited the highest prediction confidence, indicating a significant role of A4galt heterodimerization in determining enzyme specificity toward GSLs and GPs. These findings enhance our knowledge of A4galt acceptor specificity and may contribute to a better comprehension of pathomechanisms of the Shiga toxin-related diseases.
      Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
      (Copyright © 2024 Elsevier Inc. All rights reserved.)
    • Contributed Indexing:
      Keywords: A4GALT; B4GALT; Dimerization; Glycosyltransferase; Specificity
    • Accession Number:
      EC 2.4.1.- (Galactosyltransferases)
      0 (Globosides)
      0 (Glycosphingolipids)
      EC 2.4.1.- (UDP-galactose-lactosylceramide alpha 1-4-galactosyltransferase)
    • Publication Date:
      Date Created: 20240807 Date Completed: 20241108 Latest Revision: 20241108
    • Publication Date:
      20241114
    • Accession Number:
      10.1016/j.bbrc.2024.150486
    • Accession Number:
      39111055