title: Unfolding of newly made retinol-binding protein by dithiothreitol. Sensitivity to retinoids.
abstract: In cultured hepatoma HepG2 cells, serum retinol-binding protein (RBP) is secreted more rapidly in the presence of retinol than in its absence (Tosetti, F., Ferrari, N., Pfeffer, U., Brigati, C., and Vidali, G. (1992) Exp. Cell Res. 200, 467-472). In the presence of millimolar concentration of DTT, HepG2 cells synthesize fully reduced RBP within the endoplasmic reticulum (ER) which, upon removal of DTT, forms disulfide bonds post-translationally. Secretion of this post-translationally folded RBP is also dependent on the presence of retinol. Using nonreducing gel electrophoresis, we resolved disulfide-bonded RBP folding intermediates. In addition, two other intracellular folding intermediates, compact I and II, which co-migrate with mature RBP were resolved by their different sensitivity to DTT-induced unfolding. Retinol, as well as retinoic acid, stabilized both compact I and II RBP intermediates to DTT-induced unfolding, suggesting that RBP assumes different conformations in the ER in the presence and absence of a ligand. However, only RBP synthesized in the presence of retinol is rapidly secreted, indicating that the ER export quality control system recognizes RBP containing retinol, but not retinoic acid, as fully folded and competent for export. Folding of RBP so that it is stabilized to DTT reduction is not a sufficient condition for ER exit.
authors.affiliation: Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142.
authors.forename: E H
authors.initials: EH
authors.lastname: Kaji
authors.forename: H F
authors.initials: HF
authors.lastname: Lodish
mesh_terms: D004220:Disulfides / Q000737:chemistry; D004229:Dithiothreitol / Q000737:chemistry*; D006801:Humans; D010084:Oxidation-Reduction; D017510:Protein Folding; D012177:Retinol-Binding Proteins / Q000737:chemistry*; D014407:Tumor Cells, Cultured; D014801:Vitamin A / Q000737:chemistry*
chemical_list: D004220:Disulfides; D012177:Retinol-Binding Proteins; D014801:Vitamin A; D004229:Dithiothreitol