Viral protein U (Vpu) is a type-III integral membrane protein encoded by Human Immunodeficiency Virus-1 (HIV- 1). It is expressed in infected host cells and plays several roles in viral progeny escape from infected cells, including down-regulation of CD4 receptors. But key structure/function questions remain regarding the mechanisms by which the Vpu protein contributes to HIV-1 pathogenesis. Here we describe expression of Vpu in bacteria, its purification and characterization. We report the successful expression of PelB-Vpu in Escherichia coli using the leader peptide pectate lyase B (PelB) from Erwinia carotovora. The protein was detergent extractable and could be isolated in a very pure form. We demonstrate that the PelB signal peptide successfully targets Vpu to the cell membranes and inserts it as a type I membrane protein. PelB-Vpu was biophysically characterized by circular dichroism and dynamic light scattering experiments and was shown to be an excellent candidate for elucidating structural models.
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- Bacterial Expression, Correct Membrane Targeting, and Functional Folding of the HIV-1 Membrane Protein Vpu Using a Periplasmic Signal Peptide
- Deb, Arpan (Author)
- Johnson, William (Author)
- Kline, Alexander (Author)
- Scott, Boston (Author)
- Meador, Lydia (Author)
- Srinivas, Dustin (Author)
- Martin Garcia, Jose Manuel (Author)
- Dorner, Katerina (Author)
- Borges, Chad (Author)
- Misra, Rajeev (Author)
- Hogue, Brenda (Author)
- Fromme, Petra (Author)
- Mor, Tsafrir (Author)
- ASU Biodesign Center Immunotherapy, Vaccines and Virotherapy (Contributor)
- College of Liberal Arts and Sciences (Contributor)
- School of Life Sciences (Contributor)
- Biodesign Institute (Contributor)
- School of Molecular Sciences (Contributor)
- Applied Structural Discovery (Contributor)
- Personalized Diagnostics (Contributor)
- Digital object identifier: 10.1371/journal.pone.0172529
- Identifier TypeInternational standard serial numberIdentifier Value1045-3830
- Identifier TypeInternational standard serial numberIdentifier Value1939-1560
- The article is published at http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0172529
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Deb, A., Johnson, W. A., Kline, A. P., Scott, B. J., Meador, L. R., Srinivas, D., . . . Mor, T. S. (2017). Bacterial expression, correct membrane targeting and functional folding of the HIV-1 membrane protein Vpu using a periplasmic signal peptide. Plos One, 12(2). doi:10.1371/journal.pone.0172529