TABLE 1.

GOAT topology summary

L, lumenal; C, cytosolic. Plus signs indicate V5 or Myc signal level in indirect immunofluorescence. Alt glyc*, a variety of alternate glycosylation signal sequences were installed at the GOAT N terminus (Fig. 9B), and these were all lumenal. Endogenous Asn**, an endogenous Asn in a consensus NX(S/T/C) site was not glycosylated. NXS/T Mutant***, a mutation creating a consensus NX(S/T) glycosylation site was installed in the center of this loop but could not be glycosylated (Fig. 9B). Myc hGOAT, human GOAT was made with a Myc tag at this position, and results agree with the mouse GOAT result. ND, not done; GOAT with V5Glyc tag installed in positions 8a and 8b did not result in full-length protein, so positions 8c and 8d were tested with this tag. †, in a mass spectrometry experiment performed on full-length GOAT, we observed a peptide corresponding to ubiquitylation at residue Lys-141, suggesting cytosolic localization. Relative signal strength assignment shown with variable numbers of plus signs is described under “Experimental Procedures.”