The cholinesterase-like domain of thyroglobulin functions as an intramolecular chaperone
J. Clin. Invest. Jaemin Lee, et al. 118:2950 doi:10.1172/JCI35164 [
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Figure 4Efficient exit of the isolated Tg ChEL domain from the ER. 293 cells were transiently transfected with a plasmid encoding the wild-type mouse Tg ChEL domain preceded by the prolactin signal peptide (Secretory ChEL) or were untransfected (293 control). Cells were pulse labeled for 30 minutes with
35S-labeled amino acids and chased for 0 or 4 hours as indicated, at which time the cells were lysed and both lysates and media immunoprecipitated with a rabbit polyclonal anti-Tg. Immunoprecipitates from transfected cells were divided in 2 equal portions and either mock digested or digested with endoglycosidase H (Endo H). Finally, all samples were analyzed by reducing 5.5% SDS-PAGE and fluorography. The band shift observed after digestion of secretory ChEL from the 0 chase time (shift down, lane 2) is indicative of endoglycosidase H sensitivity and defines ChEL that has not yet reached the Golgi complex; in contrast, none of the secreted ChEL shows the same endoglycosidase H sensitivity, indicating intracellular transport via the Golgi complex. The lanes shown were all run on the same gel, although they are presented noncontiguously. The position of the 76-kDa molecular mass standard is shown at left.
