Abstract
Being a major metabolite for maintaining cellular homeostasis, as well as an important structural component in lipid membrane, cholesterol also plays critical roles in the life cycles of some viruses, including human immunodeficiency virus-1 (HIV-1). The involvement of cholesterol in HIV-1 infectivity, assembly and budding has made it an important research target. Viral protein R (Vpr) is an accessory protein of HIV-1, which is involved in many major events in the life cycle of HIV-1. In addition to its multi-functional roles in the HIV-1 life cycle, it is shown to interact with lipid membrane and form a cation-selective channel. In this work, we examined the effect of cholesterol on the interaction of Vpr and lipid membrane. Using calcein release assay, we found that the membrane permeability induced by the membrane binding of Vpr was significantly reduced in the presence of cholesterol in membrane. In addition, using solid-state NMR (ssNMR) spectroscopy, Vpr was shown to experience multiple chemical environments in lipid membrane, as indicated by the broad line shape of carbonyl 13C resonance of Cys-76 residue ranging from 165–178 ppm, which can be attributed to the existence of complex Vpr-membrane environments. We further showed that the presence of cholesterol in membrane will alter the distribution of Vpr in the complex membrane environments, which may explain the change of the Vpr induced membrane permeability in the presence of cholesterol.
Highlights
Study of [1-13 C, 99%] Cysteine selective labeled Viral protein R (Vpr) embedded in liposome, we showed the existence of complex Vpr-membrane environments and the distribution of Vpr among these environments was altered with the presence of cholesterol in membrane
We found the incorporation yield of Vpr in proteoliposomes made with dioleoyl-sn-glycero-3-phospho-(10 -rac-glycerol) (DOPG) lipid was in a range from 30% to 50%
No significant difference in terms of the Vpr incorporation yield was observed in cholesterol containing proteoliposomes
Summary
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. In addition to its functional roles in HIV-1 viral cycle, the interactions between Vpr and lipid membrane draw significant attention. During the HIV-1 virus particle assembly at the plasma membrane, lipid rafts containing high amount of cholesterol are targeted by Gag proteins [9,10]. We first used calcein release assay to probe the effect of cholesterol on Vpr induced membrane permeability and showed that the Vpr induced membrane permeability was reduced as the content of cholesterol in membrane was increased This result is similar to what were observed from the studies of a pore-forming peptide [35] and CPPs [36]. Even though cholesterol does not bind to Vpr nor affect the secondary structure directly, cholesterol may still modulate the Vpr-membrane interaction by changing the protein-lipid complex arrangement. We show that cholesterol, without interacting with Vpr directly, can affect the Vpr-membrane interaction
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
