Topological regulation of a transmembrane protein by luminal-to-cytosolic retrotranslocation of glycosylated sequence

Analyzing the topology of N-linked glycans by PNGase F accessibility assay

“…Their location relative to membranes cannot be determined by this assay. 1

Figure 1 Schematic illustration of the PNGase F accessibility assay The N-linked glycosylation sites and glycans are highlighted in yellow and red, respectively. Reused from Wang et al.

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“…Each of the three deglycosylation conditions reduces the apparent molecular weight of TM4SF20(C) ( Figure 2 C), as the three N-linked glycans locate at different environments: N163 at the cytosolic side, which was deglycosylated in Tube 2; N132 at the luminal side, which was deglycosylated in Tube 3 but not 2; and N148 that is inaccessible to PNGase F, which was only deglycosylated in Tube 4 when the protein was unfolded by SDS. 1 …”