analysis-protein-folding-transport-degradation-living-cells

Analysis of Protein Folding, Transport, and Degradation in Living Cells by Radioactive Pulse Chase

Radioactive pulse-chase labeling is a powerful tool for studying the conformational maturation, the transport to their functional cellular location, and ...

Utrecht University

Boston Children's Hospital

The biophysical rules that govern folding of small, single-domain proteins in dilute solutions are now quite well understood. The mechanisms underlying co-translational folding of multidomain and membrane-spanning proteins in complex cellular environments are often less clear. The endoplasmic reticulum (ER) produces a plethora of membrane and secretory proteins, which must fold and assemble correctly before ER exit - if these processes fail, misfolded species accumulate in the ER or are degraded. The ER differs from other cellular organelles in terms of the physicochemical environment and the variety of ER-specific protein modifications. Here, we review chaperone-assisted co- and post-translational folding and assembly in the ER and underline the influence of protein modifications on these processes. We emphasize how method development has helped advance the field by allowing researchers to monitor the progression of folding as it occurs inside living cells, while at the same time probing the intricate relationship between protein modifications during folding.

Co- and Post-Translational Protein Folding in the ER.

Like all other secretory proteins, the HIV-1 envelope glycoprotein gp160 is targeted to the endoplasmic reticulum (ER) by its signal peptide during synthesis. Proper gp160 folding in the ER requires core glycosylation, disulfide-bond formation and proline isomerization. Signal-peptide cleavage occurs only late after gp160 chain termination and is dependent on folding of the soluble subunit gp120 to a near-native conformation. We here detail the mechanism by which co-translational signal-peptide cleavage is prevented. Conserved residues from the signal peptide and residues downstream of the canonical cleavage site form an extended alpha-helix in the ER membrane, which covers the cleavage site, thus preventing cleavage. A point mutation in the signal peptide breaks the alpha helix allowing co-translational cleavage. We demonstrate that postponed cleavage of gp160 enhances functional folding of the molecule. The change to early cleavage results in decreased viral fitness compared to wild-type HIV.

Nicholas McCaul

Cellular Protein Chemistry

Utrecht University

Analysis of Protein Folding, Transport, and Degradation in Living Cells by Radioactive Pulse Chase

Nicholas McCaul

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Cellular Protein Chemistry

Utrecht University

Analysis of Protein Folding, Transport, and Degradation in Living Cells by Radioactive Pulse Chase

Cellular Protein Chemistry