The cholinesterase-like domain of thyroglobulin functions as an intramolecular chaperone
J. Clin. Invest. Jaemin Lee, et al. 118:2950 doi:10.1172/JCI35164 [Go to this article.]

Figure 3
Oxidation state of secreted Tg. At each chase time, cells were lysed in buffer including 20 mM N-ethylmaleimide, and the lysates and chase media immunoprecipitated with anti-Tg and analyzed by nonreducing 4% SDS-PAGE and fluorography. (A) PC Cl3 cells were pulse labeled for 10 minutes with ³⁵S-labeled amino acids and then chased for the times indicated. Immunoprecipitates were either undigested or digested with PNGase F as indicated. Folding intermediates A, B, and C, which have been characterized in previous studies (28), are shown. Also identifed are 2 closely spaced Tg disulfide isomer bands labeled D and E, respectively. Because of glycosylation differences, the mature E isoform does not comigrate with Tg secreted to the medium at 1 hour of chasing. After PNGase F digestion to remove N-glycans, all Tg forms exhibit a faster (shifted-down) mobility. Under these conditions, it is now apparent that secreted Tg comigrates with the intracellular E isomer, identifying the most oxidized band as the most mature folded form of Tg. (B) Results of an experiment identical to the one represented in A, except using recombinant Tg expressed in 293 cells, without PNGase F digestion. (C) Results of a repeat experiment of that shown in panel B, but including PNGase F digestion. The position of a 181-kDa prestained molecular weight standard is shown at left.