Glycoproteomic landscape and structural dynamics of TIM family immune checkpoints enabled by mucinase SmE

Nat Commun. 2023 Oct 4;14(1):6169. doi: 10.1038/s41467-023-41756-y.


Mucin-domain glycoproteins are densely O-glycosylated and play critical roles in a host of biological functions. In particular, the T cell immunoglobulin and mucin-domain containing family of proteins (TIM-1, -3, -4) decorate immune cells and act as key regulators in cellular immunity. However, their dense O-glycosylation remains enigmatic, primarily due to the challenges associated with studying mucin domains. Here, we demonstrate that the mucinase SmE has a unique ability to cleave at residues bearing very complex glycans. SmE enables improved mass spectrometric analysis of several mucins, including the entire TIM family. With this information in-hand, we perform molecular dynamics (MD) simulations of TIM-3 and -4 to understand how glycosylation affects structural features of these proteins. Finally, we use these models to investigate the functional relevance of glycosylation for TIM-3 function and ligand binding. Overall, we present a powerful workflow to better understand the detailed molecular structures and functions of the mucinome.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Hepatitis A Virus Cellular Receptor 2*
  • Mucins* / metabolism
  • Polysaccharide-Lyases
  • Polysaccharides / chemistry


  • hyaluronate lyase
  • Hepatitis A Virus Cellular Receptor 2
  • Mucins
  • Polysaccharide-Lyases
  • Polysaccharides