Protein folding in the endoplasmic reticulum: Lessons from the human chorionic gonadotropin β subunit

Abstract

There have been few studies of protein folding in the endoplasmic reticulum of intact mammalian cells. In the one case where the in vivo and in vitro folding pathways of a mammalian secretory protein have been compared, the folding of the human chorionic gonadotropin b subunit (hCG-b), the order of formation of the detected folding intermediates is the same. The rate and efficiency with which multidomain proteins such as hCG-P fold to native structure in intact cells is higher than in vitro, although intracellular rates of folding of the b subunit can be approached in vitro in the presence of an optimal redox potential and protein disulfide isomerase. Understanding how proteins fold in vivo may provide a new way to diagnose and treat human illnesses that occur due to folding defects.Keywords: disease states related to protein misfolding; hCG-P subunit as a model of secretory protein folding; molecular chaperones; protein misfolding; role of disulfide bonds and N-linked glycosylation in protein folding, assembly, and secretion Protein folding of secreted proteinsA growing number of human diseases are now known to be related to protein folding defects, and thus it is important to understand the rules that govern protein folding in vivo. The focus of this review is the mechanisms that regulate protein folding in intact cells, particularly those proteins synthesized in the endoplasmic reticulum (ER) of mammalian cells and targeted for translocation to the cell surface or for secretion.Many of the eukaryotic proteins whose folding and assembly have been studied in vivo are membrane or secreted proteins. They follow a similar route to the cell surface. (1) Synthesis occurs in the rough ER. (2) Nascent proteins are translocated into the cisternal space of the ER where the signal peptide is cleaved; initial co-translational folding involving secondary structure and some native tertiary structure occurs; addition of high-mannose N-linked oligosaccharides and initial processing of N-linked oligosaccharide chains (for glycoproteins) takes place; formation of disulfide bonds occurs, and for multimeric proteins, oligomerization or subunit assembly is attained along with achievement of native structure. (3) The proteins destined for the cell surface or secretion are translocated to the Golgi apparatus, further processed, and then either translocated to the cell surface or packaged into secretory vesicles for secretion.There are some similarities as well as differences between intracellular protein folding and protein folding in test tubes. For instance, for the tailspike protein of Salmonella typhimuriurn phage P22 (Fuchs et al., 1991;Mitraki et al., 1991) and the human chorionic gonadotropin b subunit (hCG-b) , the intermediates in the folding pathway for proteins appear to be the same in vivo and in vitro, but the rate and efficiency with which proteins achieve final native state in vivo is higher than in vitro. There are also many observations both in vivo and in vitro showing that correct folding is in competition...

Alternatively Folded Choriogonadotropin Analogs

“…This result is consistent with the model of hCG assembly proposed by Ruddon et al (17) and shows that the conformation of the seatbelt is not restricted to the position it occupies in hCG (2, 3). These data also support a prediction TABLE IV Binding of selected analogs in alternate sandwich assays Analogs were captured with A113, an antibody to the ␣-subunit, and detected with radiolabeled antibodies to the ␤-subunit.…”

“…This suggests that the amino-terminal half of the seatbelt is not as mobile as the carboxyl-terminal half or that the amino-terminal half of the seatbelt contributes to interactions between the subunits needed for docking when the seatbelt is not latched. This observation is also consistent with the proposal that the small seatbelt loop is formed prior to docking of the subunits (17).…”

Human Lutropin (hLH) and Choriogonadotropin (CG) Are Assembled by Different Pathways

“…The C terminus of the seatbelt is "latched" by a disulfide bond to a cysteine in loop 1 of the ␤-subunit. Earlier studies suggested that heterodimer assembly occurred by a "wraparound" mechanism in which the seatbelt became latched after the subunits docked (3). Contrary to these studies, we reported that the assembly of human choriogonadotropin (hCG), human follitropin (hFSH), and human thyrotropin (hTSH) occurred in the endoplasmic reticulum only after the seatbelt was latched (4,5).…”

Pathogenesis of Alzheimer's Disease: Role of Amyloid-beta and Hyperphosphorylated Tau Protein

“…On the other hand some diseases are due to the synthesis of a mutant or misfolded protein, which gets a novel activity (such as the tendency to from aggregates) with pathological characteristics. Different human diseases, which involve protein misfolding have been studied [60,61] .…”