Physico-Chemical Mechanisms of the Functioning of Membrane-Active Proteins of Enveloped Viruses.

Over the past few years, the attention of the whole world has been riveted to the emergence of new dangerous strains of viruses, among which a special place is occupied by coronaviruses that have overcome the interspecies barrier in the past 20 years: SARS viruses (SARS), Middle East respiratory syndrome (MERS), as well as a new coronavirus infection (SARS-CoV-2), which caused the largest pandemic since the Spanish flu in 1918. Coronaviruses are members of a class of enveloped viruses that have a lipoprotein envelope. This class also includes such serious pathogens as human immunodeficiency virus (HIV), hepatitis, Ebola virus, influenza, etc. Despite significant differences in the clinical picture of the course of disease caused by enveloped viruses, they themselves have a number of characteristic features, which determine their commonality. Regardless of the way of penetration into the cell-by endocytosis or direct fusion with the cell membrane-enveloped viruses are characterized by the following stages of interaction with the target cell: binding to receptors on the cell surface, interaction of the surface glycoproteins of the virus with the membrane structures of the infected cell, fusion of the lipid envelope of the virion with plasma or endosomal membrane, destruction of the protein capsid and its dissociation from the viral nucleoprotein. Subsequently, within the infected cell, the newly synthesized viral proteins must self-assemble on various membrane structures to form a progeny virion. Thus, both the initial stages of viral infection and the assembly and release of new viral particles are associated with the activity of viral proteins in relation to the cell membrane and its organelles. This review is devoted to the analysis of physicochemical mechanisms of functioning of the main structural proteins of a number of enveloped viruses in order to identify possible strategies for the membrane activity of such proteins at various stages of viral infection of the cell.

New Insights into the Interaction of Class II Dihydroorotate Dehydrogenases with Ubiquinone in Lipid Bilayers as a Function of Lipid Composition.

The fourth enzymatic reaction in the de novo pyrimidine biosynthesis, the oxidation of dihydroorotate to orotate, is catalyzed by dihydroorotate dehydrogenase (DHODH). Enzymes belonging to the DHODH Class II are membrane-bound proteins that use ubiquinones as their electron acceptors. We have designed this study to understand the interaction of an N-terminally truncated human DHODH (HsΔ29DHODH) and the DHODH from Escherichia coli (EcDHODH) with ubiquinone (Q10) in supported lipid membranes using neutron reflectometry (NR). NR has allowed us to determine in situ, under solution conditions, how the enzymes bind to lipid membranes and to unambiguously resolve the location of Q10. Q10 is exclusively located at the center of all of the lipid bilayers investigated, and upon binding, both of the DHODHs penetrate into the hydrophobic region of the outer lipid leaflet towards the Q10. We therefore show that the interaction between the soluble enzymes and the membrane-embedded Q10 is mediated by enzyme penetration. We can also show that EcDHODH binds more efficiently to the surface of simple bilayers consisting of 1-palmitoyl, 2-oleoyl phosphatidylcholine, and tetraoleoyl cardiolipin than HsΔ29DHODH, but does not penetrate into the lipids to the same degree. Our results also highlight the importance of Q10, as well as lipid composition, on enzyme binding.

Structure, interactions and membrane topology of HIV gp41 ectodomain sequences.

The gp41 type I membrane protein is part of the trimeric Env complex forming the spikes at the HIV surface. By interacting with cellular receptors, the Env protein complex initiates the infectious cycle of HIV. After the first contact has been established Env disassembles by shedding gp120 while the remaining gp41 undergoes a number of conformational changes which drive fusion of the cellular and the viral membranes. Here we investigated the membrane interactions and oligomerization of the two gp41 heptad repeat domains NHR and CHR. While these are thought to form a six-helix bundle in the post-fusion state little is known about their structure and role during prior fusion events. When investigated in aqueous buffer by CD and fluorescence quenching techniques the formation of NHR/CHR hetero-oligomers is detected. An equilibrium of monomers and hetero-oligomers is also observed in membrane environments. Furthermore, the partitioning to POPC or POPC/POPG 3/1 vesicles of the two domains alone or in combination has been studied. The membrane interactions were further characterized by 15N solid-state NMR spectroscopy of uniaxially oriented samples which shows that the polypeptide helices are oriented parallel to the bilayer surface. The 31P solid-state NMR spectra of the same samples are indicative of considerable disordering of the membrane packing. The data support models where NHR and CHR insert in the viral and cellular membranes, respectively, where they exhibit an active role in the membrane fusion events.