{Reference Type}: Journal Article
{Title}: Exposure to brefeldin A induces unusual expression of hybrid- and complex-type free N-glycans in HepG2 cells.
{Author}: Sugiura K;Kawai Y;Yamamoto A;Yoshioka H;Kiyohara Y;Iida A;Ozawa Y;Nishikawa M;Miura N;Hanamatsu H;Furukawa JI;Shinohara Y;
{Journal}: Biochim Biophys Acta Gen Subj
{Volume}: 1867
{Issue}: 5
{Year}: 05 2023
{Factor}: 4.117
{DOI}: 10.1016/j.bbagen.2023.130331
{Abstract}: This study determined the effect of brefeldin A (BFA) on the free N-glycomic profile of HepG2 cells to better understand the effect of blocking intracellular vesicle formation and transport of proteins from the endoplasmic reticulum to the Golgi apparatus. A series of exoglycosidase- and endoglycosidase-assisted analyses clarified the complex nature of altered glycomic profiles. A key feature of BFA-mediated alterations in Gn2-type glycans was the expression of unusual hybrid-, monoantennary- and complex-type free N-glycans (FNGs). BFA-mediated alterations in Gn1-type glycans were characterized by the expression of unusual hybrid- and monoantennary-FNGs, without significant expression of complex-type FNGs. A time course analysis revealed that sialylated hybrid- and complex-type Gn2-type FNGs were generated later than asialo-Gn2-type FNGs, and the expression profiles of Gn2-type FNGs and N-glycans were found to be similar, suggesting that the metabolic flux of FNGs is the same as that of protein-bound N-glycans. Subcellular glycomic analysis revealed that almost all FNGs were detected in the cytoplasmic extracts. Our data suggest that hybrid-, monoantennary- and complex-type Gn2-type FNGs were cleaved from glycoproteins in the cytosol by cytosolic PNGase, and subsequently digested by cytosolic endo-β-N-acetylglucosaminidase (ENGase) to generate Gn1-type FNGs. The substrate specificity of ENGase explains the limited expression of complex Gn1 type FNGs.