Insights into the structure-function relationship of the HIV-1 Vpu protein.

Abstract

HIV-1 is the causative agent of HIV/AIDS. Annually, hundreds of thousands of people carrying the virus die from virus-related causes. Still, the means to cure the HIV-caused diseases are limited. Acquiring detailed knowledge about the viral mechanisms to modulate cellular processes is vastly needed for the development of efficient anti-viral treatments. We study the molecular mechanisms of HIV-1-host interactions, particularly the virus-encoded proteins with major roles in the virus lifecycle. The focus is on the Vpu protein, which forms ion-conducting homo-oligomers in cellular membranes but the protein monomer is also deemed physiologically relevant due its interaction with host proteins. However, the structure of full-length (FL) Vpu, required for structure-based drug design, has not been resolved. Recently, we made significant progress toward the understanding of FL Vpu quaternary structure. We used negative staining electron microscopy (nsEM) and size exclusion chromatography to characterize Vpu oligomerization. We generated a chimera protein of maltose binding protein fused to Vpu N-terminal (MBP-Vpu) to increase the protein size to aid EM experiments. We found that the MBP-Vpu forms a stable oligomer (likely pentamer) in solution, possibly driven by the hydrophobic effect and TM self-association. We further found that when transferred to the lipid environment, these oligomers restructured significantly: along with the pentamers, low-order oligomers were observed as well, or under specific lipid conditions in lyso lipid Vpu formed liner array-like aggregates. This emphasizes the role of lipid environment on Vpu oligomer organization. All these findings are novel and the observed Vpu quaternary structures are most likely physiologically relevant. Our ongoing cryo-EM experiments aim to solve the high-resolution structure of Vpu oligomer. The characterization of Vpu-drug binding is ongoing in our lab as well.