Structure and topology around the cleavage site regulate post-translational cleavage of the HIV-1 gp160 signal peptide
Abstract
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 that 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.
Article and author information
Author details
Funding
Nederlandse Organisatie voor Wetenschappelijk Onderzoek
- Nicholas McCaul
- Matthias Quandte
- Aafke Land
- Ineke Braakman
Netherlands AIDS Fund
- Aafke Land
Seventh Framework Programme (ITN 'Virus Entry')
- Nicholas McCaul
- Matthias Quandte
- Ineke Braakman
National Institutes of Health (NIH AI-51519)
- Erik Lee Snapp
Swedish Cancer Foundation
- IngMarie Nilsson
- Gunnar von Heijne
Knut and Alice Wallenberg Foundation
- Gunnar von Heijne
European Research Council (ERC-StG-2011-280829-SHEV)
- Rogier W Sanders
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2017, Snapp et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 5,151
- views
-
- 580
- downloads
-
- 43
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Further reading
-
- Biochemistry and Chemical Biology
Class-B1 G-protein-coupled receptors (GPCRs) are an important family of clinically relevant drug targets that remain difficult to investigate via high-throughput screening and in animal models. Here, we engineered PAClight1P78A, a novel genetically encoded sensor based on a class-B1 GPCR (the human PAC1 receptor, hmPAC1R) endowed with high dynamic range (ΔF/F0 = 1100%), excellent ligand selectivity, and rapid activation kinetics (τON = 1.15 s). To showcase the utility of this tool for in vitro applications, we thoroughly characterized and compared its expression, brightness and performance between PAClight1P78A-transfected and stably expressing cells. Demonstrating its use in animal models, we show robust expression and fluorescence responses upon exogenous ligand application ex vivo and in vivo in mice, as well as in living zebrafish larvae. Thus, the new GPCR-based sensor can be used for a wide range of applications across the life sciences empowering both basic research and drug development efforts.
-
- Biochemistry and Chemical Biology
- Structural Biology and Molecular Biophysics
Metal-ion-dependent nucleases play crucial roles in cellular defense and biotechnological applications. Time-resolved crystallography has resolved catalytic details of metal-ion-dependent DNA hydrolysis and synthesis, uncovering the essential roles of multiple metal ions during catalysis. The histidine-metal (His-Me) superfamily nucleases are renowned for binding one divalent metal ion and requiring a conserved histidine to promote catalysis. Many His-Me family nucleases, including homing endonucleases and Cas9 nuclease, have been adapted for biotechnological and biomedical applications. However, it remains unclear how the single metal ion in His-Me nucleases, together with the histidine, promotes water deprotonation, nucleophilic attack, and phosphodiester bond breakage. By observing DNA hydrolysis in crystallo with His-Me I-PpoI nuclease as a model system, we proved that only one divalent metal ion is required during its catalysis. Moreover, we uncovered several possible deprotonation pathways for the nucleophilic water. Interestingly, binding of the single metal ion and water deprotonation are concerted during catalysis. Our results reveal catalytic details of His-Me nucleases, which is distinct from multi-metal-ion-dependent DNA polymerases and nucleases.