Research Article: A single β-octyl glucoside molecule induces HIV-1 Nef dimer formation in the absence of partner protein binding

Date Published: February 7, 2018

Publisher: Public Library of Science

Author(s): Mousheng Wu, John J. Alvarado, Corinne E. Augelli-Szafran, Roger G. Ptak, Thomas E. Smithgall, Wenqing Xu.

http://doi.org/10.1371/journal.pone.0192512

Abstract

The HIV-1 Nef accessory protein is essential for viral pathogenicity and AIDS progression. Nef forms complexes with multiple host cell factors to facilitate viral replication and promote immune escape of HIV-infected cells. Previous X-ray crystal structures demonstrate that Nef forms homodimers, the orientation of which are influenced by host cell binding partners. In cell-based fluorescence complementation assays, Nef forms homodimers at the plasma membrane. However, recombinant Nef proteins often exist as monomers in solution, suggesting that membrane interaction may also trigger monomer to dimer transitions. In this study, we show that monomeric Nef core proteins can be induced to form dimers in the presence of low concentrations of the non-ionic surfactant, β-octyl glucoside (βOG). X-ray crystallography revealed that a single βOG molecule is present in the Nef dimer, with the 8-carbon acyl chain of the ligand binding to a hydrophobic pocket formed by the dimer interface. This Nef-βOG dimer interface involves helix αB, as observed in previous dimer structures, as well as a helix formed by N-terminal residues 54–66. Nef dimer formation is stabilized in solution by the addition of βOG, providing biochemical validation for the crystal structure. These observations together suggest that the interaction with host cell lipid mediators or other hydrophobic ligands may play a role in Nef dimerization, which has been previously linked to multiple Nef functions including host cell protein kinase activation, CD4 downregulation, and enhancement of HIV-1 replication.

Partial Text

The nef genes of human and simian immunodeficiency viruses (HIV-1, HIV-2, and SIV) encode accessory proteins of 27–35 kDa critical for viral pathogenesis [1,2]. While Nef is not essential for HIV-1 replication in vitro, it enhances virus replication in vivo and promotes AIDS progression. Deletion of Nef attenuates SIV replication, pathogenicity and AIDS progression in non-human primates [3]. More recent studies have shown that Nef is also required for HIV-1 replication and CD4+ T cell loss in humanized mice [4,5]. Conversely, targeted expression of Nef alone to the CD4+ cellular compartment is sufficient to induce an AIDS-like syndrome in transgenic mice [6]. Consistent with these observations, some individuals infected with Nef-defective HIV-1 remain asymptomatic for many years in the absence of antiretroviral therapy [7,8]. These studies demonstrate the importance of Nef to HIV-1 pathogenesis in vivo, and support the development of drugs targeting this virulence factor as a new approach to HIV-1 therapy [9].

The structure of Nef (51–205) reported here represents a novel homodimer conformation that is induced by interaction with a single molecule of βOG. Many of the residues involved in the dimer interface and βOG-binding pocket are conserved across Nef alleles derived from multiple HIV-1 subtypes as well as SIV, suggesting that most Nef proteins have the potential to adopt the βOG -induced dimer structure. The finding that the dimer is stabilized by the hydrophobic tail of βOG both in the crystal and in solution raises the possibility that host cell lipids may also affect Nef dimerization, including the myristate modifications present on several Nef binding partners as well as Nef itself. Our results also highlight the remarkably dynamic capacity of HIV-1 Nef to form multiple quaternary structures, despite the highly conserved nature of its folded core region. Identification of small molecules that occupy the βOG-binding pocket reported here may restrict Nef dynamics and act as global inhibitors of the multiple functions of this critical HIV-1 virulence factor.

 

Source:

http://doi.org/10.1371/journal.pone.0192512

 

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