Poster Presentation HUPO 2019 - 18th Human Proteome Organization World Congress

Suppression of terminal modifications of N-glycans by bisecting GlcNAc (#621)

Yasuhiko Kizuka 1 , Miyako Nakano 2 , Naoyuki Taniguchi 3
  1. Gifu University, Gifu, GIFU, Japan
  2. Hiroshima University, Hiroshima, Japan
  3. Osaka International Cancer Institute, Osaka

Bisecting GlcNAc, a branched GlcNAc in N-glycan synthesized by GnT-III (encoded by MGAT3 gene), is most highly expressed in brain. Our previous studies using Mgat3-deficient mice showed that bisecting GlcNAc promotes Alzheimer’s pathology by positively regulating the functions of amyloid beta-producing enzyme BACE1. However, physiological functions of bisecting GlcNAc remain largely unclear.

Several in vitro biochemical studies already showed that introduction of bisecting GlcNAc suppresses further actions of other GlcNAc-branching enzymes, such as GnT-IV and -V, suggesting that bisecting GlcNAc regulates overall N-glycan profiles in vivo. To clarify this issue, we analyzed N-glycan structures of Mgat3-deficient brain and found that various terminal modifications were upregulated in Mgat3-knockout, such as Lewis-fucose, sialic acid and HNK-1 epitopes. The levels of glycosyltransferase mRNAs and their donor substrates were unchanged between WT and Mgat3-knockout. In contrast, our enzyme assays using bisected and non-bisected acceptor substrates revealed that most enzymes working on N-glycan terminals prefer the non-bisected glycan as a substrate, indicating that the upregulation of terminal N-glycan epitopes were attributed tosubstrate specificity of glycosyltransferases. Molecular dynamics simulation of glycosyltransferase-glycan complexes showed that the presence of bisecting GlcNAc changed overall N-glycan conformation from extended type to back-fold type where alpha1,6-arm loses interaction with the enzymes, suggesting that the bisected glycan is not preferred by various glycosyltransferases for N-glycan terminals due to alteration of N-glycan conformation. These results highlight the roles of bisecting GlcNAc as a general suppressor of terminal modifications of N-glycans, providing new insight into how protein N-glycosylation is regulated in cells.