Virus Spike Protein – an overview
Viral Spike protein (S) binds to the ACE2 receptor, followed by viral fusion with the cell membrane, which involves priming of the S protein by proteases on the cell surface, and culminates in the entry and replication of the virus in the host cell cytoplasm [24].
Natural protein fibres
3.2 Viral Spike Protein
The structural characteristics of virus coats (capsids) are highly relevant to virus propagation, and are therefore the subject of intensive study. The coat must contain and protect the nucleic acid (genetic) contents, be resilient against impact, be capable of broaching the outer wall of a target cell, and provide a secure pathway for conducting nucleic acid into the target.
Hollow spikes on the capsid fulfill the latter two roles, from which it has been deduced that they must have unusually high strength and stiffness in axial compression. Because compressive strength and stiffness have been a long-term but elusive goal of polymer science, the hierarchical structure of spike proteins deserves careful attention.
(a) Cross-β-sheets
Viral spike protein contains several repeats of relatively short β-strand-forming amino acid sequences. The chain folds back and forth to assemble a β-sheet, stabilized by intramolecular hydrogen bonds (Fig. 2). Because the long axis of the sheet is transverse rather than parallel to the molecular backbone, the structure in this case is known as a cross-β-sheet. Some silks, e.g., the egg stalks of green lacewing flies, are also reinforced by cross-β-sheet crystals.
(b) Higher-order structure
The structure varies significantly between different virus types. For both rotavirus and human adenovirus, there is evidence that three cross-β-sheets interact to form a hollow trimeric box beam that resists buckling in compression. Structural characterization is hampered by the small size of individual spikes (length⩽30 nm and width⩽5 nm). Hydrophobic bonding is presumed to be important in stabilizing their structure, since 70% of the amino acid residues are hydrophobic in representative cases.
Genetically engineered model polymers, based on multiple consecutive copies of the principal repeated sequence in native adenovirus spike protein, can self-assemble into a liquid crystalline phase in solution. However, trimeric box beams have not been found in the hierarchical microstructure of fibers spun from this phase. The spun fibers are formed under significantly off-equilibrium conditions, and should not be expected to have the same internal structure as the native spikes.
(c) Properties
In tensile tests conducted on dry fiber, the breaking strength is ∼0.3 GPa, the stiffness ∼4 GPa, and the elongation to failure ∼ 30%. Given the evidence that the native spikes rely on hydrophobic bonding to maintain their structure, the properties of analog fibers as measured in air are likely to be inferior compared to results obtained in water—even if the detailed microstructure of the native material could be reproduced. In other words, the natural material is designed to work in an aqueous medium, and attempts to mimic its properties must take this reality into account.
Animal Rhabdoviruses
Virion Properties
Morphology
Rhabdovirus virions are 100–400 nm long and 50–100 nm in diameter (Figure 1). Viruses appear bullet-shaped. From the outer to the inner side of the virion, one can distinguish the envelope covered with viral glycoprotein spikes and, internally, the nucleocapsid with helical symmetry consisting of the nucleoprotein tightly bound to genomic RNA.
Genome Organization and Genetics
All rhabdoviruses contain a genome consisting of a nonsegmented single-stranded negative-sense RNA molecule with a size in the range of approximately 8.9–15 kbp. This RNA molecule contains at least five open reading frames (ORFs) encoding five virion proteins in the order (3′–5′): nucleoprotein (N); phosphoprotein (P); matrix protein (M); glycoprotein (G); and polymerase (L). Viruses in the genus Ephemerovirus contain several additional ORFs between the G and L genes, which encode a second glycoprotein (GNS) and several other nonstructural proteins.
Similarly, in the genus Novirhabdovirus, a sixth functional cistron between the G and L genes encodes a nonstructural protein (NV) of unknown function. The unclassified rhabdoviruses, sigma virus infecting flies (Drosophila spp.) and plant rhabdoviruses in the genera Cytorhabdovirus and Nucleorhabdovirus also contain an additional ORF which is located between the P and M genes. Flanders virus from mosquitoes (Culista melanura) has a complex arrangement of genes and pseudogenes in the same genome region.
Nucleotide sequence analysis of Tupaia virus from the tree shrew (Tipiai belangeri) has identified an additional gene encoding a small hydrophobic protein between the M and G genes, and genome sequence analysis of Wongabel virus, an unassigned rhabdovirus isolated from biting midges (Culicoides austropalpalis), has revealed that it contains five additional genes that appear to be novel.
The function of these other proteins (including additional glycoproteins) is not yet known. Therefore, despite preservation of a characteristic particle morphology, the Rhabdoviridae includes viruses that display a wide genetic diversity (Figure 2).
Relatively low sequence identities across the Rhabdoviridae prevent the construction of a family phylogeny. One approach to determining the phylogenetic relationships among the rhabdoviruses, as well as the identification of new viral species, is to utilize the conserved regions that have been identified in alignments of the N and L genes.
Herpesviruses
Family Herpesviridae
The family Herpesviridae is a family of large DNA viruses containing over 100 different virus species that infect hosts ranging from humans to birds to reptiles. Classification of a virus as a member of the family Herpesviridae is based on a shared virion structure: a linear, double-stranded DNA genome is contained within a central core, surrounded by an icosahedral capsid.
This capsid is in turn surrounded first by an amorphous protein layer, known as the tegument, and then by an envelope containing viral glycoprotein spikes (Figure 1). Herpesviruses also share four significant biological properties:
- 1.They encode a large number of enzymes involved in nucleic acid metabolism, DNA synthesis, and processing of proteins.
- 2.The synthesis of viral DNAs and caspid assembly occurs in the nucleus of the infected cell. During infection, virus-specific compartments are assembled within the nucleus of the infected cell, commonly referred to as replication compartments (Figure 2). It is within these compartments that viral DNA replication, late viral gene expression, and encapsidation of progeny viral genomes occur. These compartments lead to the formation of basophilic nuclear inclusion bodies, which are diagnostic of herpes virus infection.
- 3.Production of infectious progeny virus is generally accompanied by the destruction of the infected cell.
- 4.The viruses are able to establish a latent infection in their natural hosts.
There are currently eight herpesviruses that are known to infect humans: herpes simplex virus (HSV)-1 and HSV-2, human cytomegalovirus (HCMV), varicella zoster virus (VZV), Epstein–Barr virus (EBV), Kaposi’s sarcoma-associated herpesvirus (KSHV) and human herpesvirus (HHV)-6 and HHV-7. These viruses, along with the majority of herpesviruses that infect other mammals and birds, have been divided into three subfamilies, alpha, beta, and gamma, based on the biological properties of the viruses.
HSV-1, HSV-2, and VZV are members of the Alphaherpesvirinae subfamily, EBV and KSHV are both members of the Gammaherpesvirinae subfamily, while the remaining viruses, HCMV and HHV-6A, HHV-6B, and HHV-7 are all members of the Betaherpesvirinae subfamily. Despite the many similarities in structure and biological properties shared by herpesviruses, it is not surprising that in a group of this size there are also many differences.
Host range, length of replicative cycle, cell type in which latency is established, and clinical manifestations of disease all vary among the different members of the family.
Influenza
Causative Agent
Human influenza is caused by influenza A and B viruses. Although there is no antibody cross-reactivity between the proteins encoded by these two types of influenza viruses, their replication cycle is very similar. Both are negative-strand RNA virus belonging to the orthomyxovirus group whose genome is composed of eight RNA segments, each segment encoding one or two viral proteins. The virions are enveloped, with two types of viral glycoproteins (spikes) inserted in the envelope: the hemagglutinin (HA) and neuraminidase (NA).
The HA recognizes the viral receptor, sialic acid-containing molecules, and therefore is responsible for the attachment of the virus to cells. This results in the internalization of the virus into an endosome, where acidification of the pH causes a conformational change of the HA that triggers fusion of viral envelope with the membrane of the endosome, resulting in the injection of the viral genome into the cytoplasm.
he viral genomic RNAs are encapsidated by the viral nucleoprotein (NP), in the form of ribonucleoprotein (RNP). The viral RNPs have also bound the viral RNA-dependent RNA polymerase, a complex of three protein subunits, PB2, PB1, and PA. The RNPs are surrounded by a layer of viral matrix protein (M1), but this layer becomes dissociated (uncoating) from the RNPs by its previous exposure to acidic pH just prior to the fusion event.
This exposure is mediated by a small viral ion channel or M2 protein (BM2 in the case of influenza B virus) that like the HA and NA is anchored into the viral envelope. The M2 protein transports protons from the acidified endosome to the interior of the virion, resulting in dissociation of the M1 from the viral RNPs. Uncoated RNPs are transported to the nucleus where replication and transcription takes place through the activity of the viral RNA polymerase, which synthesizes a replicative positive-strand intermediate as well as mRNAs.
Among the newly synthesized viral proteins, the viral nuclear export protein, or NEP, is responsible for the exit of the newly synthesized viral RNPs from the nucleus back to the cytoplasm, and budding at the plasma membrane results in the formation of new enveloped virions that spread to new cells.
The NA is required for efficient spreading by removing sialic acids present in newly synthesized virions, which otherwise will be bound by HAs from adjacent virions, resulting in viral clumping. Both influenza A and B virus synthesize a viral nonstructural protein or NS1 in infected cells that subverts the cellular antiviral response. In addition, most of the strains of influenza A virus encode a short nonstructural viral polypeptide, PB1-F2, that localizes to the mitochondria and modulates the apoptotic cellular pathways.
Antibody-dependant cellular Phagocytosis and its impact on pathogen control
Targeting of Viruses to FcRs
Viral infection of target cells in the absence of virus-specific antibodies is in general dependent on the interaction between viral spike proteins and corresponding receptors on target cells, thereby conferring a specific tropism of individual viruses to their target cells. Antibody-opsonized virus particles, however, may gain access to additional FcR-expressing target cells. Uptake of opsonized virus particles into FcR-bearing phagocytes via FcR-dependent phagocytosis may have different outcomes for the infection process and for the ensuing spread and control of the virus infection.
While very little is known about the exact intracellular fate of opsonized virus particles and how this relates to their infectivity in phagocytes, most available data indicate that FcR-mediated uptake into phagocytes does not interfere with intracellular viral replication. However, in situations when opsonizing antibodies have direct neutralizing effects on the fusion or uncoating process of the virus, and when this property is maintained under acidic conditions as present in late phagosomes/lysosomes,106–108 FcR-mediated uptake of opsonized viruses might restrict replication within phagocytes.
In cases where the infected phagocyte does not support the requirements of a specific viral life cycle, FcR-mediated uptake of opsonized viruses may also lead to enhanced control of viral replication.
Arenaviridae*
Virus Replication
Arenaviruses replicate noncytolytically to high titer in many kinds of cells, including Vero (African green monkey) and BHK-21 (baby hamster kidney) cells. Virus replication occurs in the cytoplasm. The viral spike glycoprotein attaches to a cell receptor, which can be transferrin receptor 1 for several New World arenaviruses and alpha-dystroglycan for lymphocytic choriomeningitis virus. Entry and endosomal uptake occur via either clathrin-dependent or clathrin-independent pathways, perhaps depending on the individual species of arenavirus.
Specifically, it is proposed that New World arenaviruses such as Junin virus utilize clathrin-mediated endocytosis, whereas Old World arenaviruses such as lymphocytic choriomeningitis and Lassa viruses utilize a clathrin-independent pathway. Because the genome of the single-stranded, negative-sense RNA viruses cannot be translated directly, the first step in replication is activation of the virion RNA polymerase (transcriptase).
The ambisense coding strategy of the arenavirus genome means that only the nucleoprotein (N) and polymerase (L protein) mRNAs are transcribed directly from genomic RNA before translation (Fig. 23.2). Newly synthesized polymerase and nucleocapsid proteins facilitate the synthesis of full-length, complementary-sense RNA, which then serves as template for the transcription of glycoprotein (GP) and zinc-binding protein (Z) mRNAs and the synthesis of more full-length, negative-sense RNA.
Budding of virions occurs from the plasma membrane (Fig. 23.3). Arenaviruses have limited ability to lyse the cells in which they replicate, usually producing a carrier state in which defective-interfering particles are produced (see Chapter 2: Virus Replication).
After an initial period of active virus transcription, translation, genome replication, and production of progeny virions, viral gene expression is downregulated, and cells enter a state of persistent infection wherein virion production continues for an indefinite period but at a greatly reduced rate providing a mechanism for persistent nonlethal infection of potential reservoir hosts (Table 23.2).
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