Bacterial expression, correct membrane targeting and functional folding of the HIV-1 membrane protein Vpu using a periplasmic signal peptide.

• Main Authors: Arpan Deb, William A Johnson, Alexander P Kline, Boston J Scott, Lydia R Meador, Dustin Srinivas, Jose M Martin-Garcia, Katerina Dörner, Chad R Borges, Rajeev Misra, Brenda G Hogue, Petra Fromme, Tsafrir S Mor
• Format: Article
• Language: English
• Published: Public Library of Science (PLoS) 2017-01-01
• Series: PLoS ONE
• Online Access: http://europepmc.org/articles/PMC5321405?pdf=render

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.