Chemical modification of hyaluronan oligosaccharides differentially modulates hyaluronan–hyaladherin interactions

Rebecca J. Dodd, Charles D. Blundell, Benedict M. Sattelle, Jan J. Enghild, Caroline M. Milner, Anthony J. Day*

*Corresponding author for this work

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

Abstract

The glycosaminoglycan hyaluronan (HA) is a ubiquitous, nonsulfated polysaccharide with diverse biological roles mediated through its interactions with HA-binding proteins (HABPs). Most HABPs belong to the Link module superfamily, including the major HA receptor, CD44, and secreted protein TSG-6, which catalyzes the covalent transfer of heavy chains from inter-α-inhibitor onto HA. The structures of the HA-binding domains (HABDs) of CD44 (HABD_CD44) and TSG-6 (Link_TSG6) have been determined and their interactions with HA extensively characterized. The mechanisms of binding are different, with Link_TSG6 interacting with HA primarily via ionic and CH−π interactions, whereas HABD_CD44 binds solely via hydrogen bonds and van der Waals forces. Here, we exploit these differences to generate HA oligosaccharides, chemically modified at their reducing ends, that bind specifically and differentially to these target HABPs. Hexasaccharides (HA6AN) modified with 2- or 3-aminobenzoic acid (HA6-2AA, HA6-3AA) or 2-amino-4-methoxybenzoic acid (HA6-2A4MBA), had increased affinities for Link_TSG6 compared to unmodified HA6AN. These modifications did not increase the affinity for CD44_HABD. A model of HA6-2AA (derived from the solution dynamic 3D structure of HA4-2AA) was docked into the Link_TSG6 structure, providing evidence that the 2AA-carboxyl forms a salt bridge with Arginine-81. These modeling results informed a second series of chemical modifications for HA oligosaccharides, which again showed differential binding to the two proteins. Several modifications to HA4 and HA6 were found to convert the oligosaccharide into substrates for heavy chain transfer, whereas unmodified HA4 and HA6 are not. This study has generated valuable research tools to further understand HA biology.

Original languageEnglish
Article number107668
JournalThe Journal of Biological Chemistry
Volume300
Issue9
ISSN0021-9258
DOIs
Publication statusPublished - Sept 2024

Keywords

  • CD44
  • chemical modification
  • extracellular matrix
  • hyaluronan
  • hyaluronan–protein interactions
  • structural biology
  • thermodynamics
  • TSG-6

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