FIGURE 5.

The V5Glyc tag is a novel dual purpose topology reporter. The V5Glyc tag is a neutral, hairpin-like tag containing an optimized 8-residue consensus for N-linked glycosylation, sandwiched between two V5 epitope tags. This tag was designed using the NetNGlyc server⁸ to provide a consistent context that can be glycosylated if located in the ER lumen. Here, we show a hypothetical membrane protein with asparagine residues in consensus sequences (i.e. NXS or NXT motifs) on both sides of the membrane. Only lumenal asparagines in the appropriate context and spaced far enough from the membrane can be N-glycosylated.