Virion structural proteins

The retrovirus genome has three regions, gag, pol, and env, that encode proteins, and all of these proteins are found in the virion. The gag region encodes a polyprotein that is cleaved to form internal virion structural proteins. The pol... 

Protease is responsible for cleaving these precursor molecules to produce the final structural proteins of the mature virion core. By preventing post-translational cleavage of the Gag-Pol polyprotein, protease inhibitors (Pis) prevent the processing of viral proteins into functional conformations, resulting in the production of immature, noninfectious viral particles (Figure 49-4). Protease inhibitors are active against both HIV-1 and HIV-2 unlike the NRTTs, however, they do not need intracellular activation. 

AIDS virion structure. Numbers associated with proteins are kilodalton masses (e.g., gpl20 is a 120-kd protein). ( Scientific American, Inc., George V. Kelvin. Reprinted by permission.)... 

Fig. 14.1 The life cycle of coxsackievirus B3. CVB3 starts its life cycle by attaching to its receptor CAR and coreceptor DAF. Internalized virus releases its viral RNA, which can be used as the template for translation of polyprotein or transcription by RNA-dependent RNA polymerase 3D to replicate its genome. The polyprotein is self-cleaved by virus-encoded proteases to release structural proteins and nonstructural proteins. Later, structural proteins and viral RNA will assemble into progeny virions to be released from infected cell. Abbreviations CVB3, coxsackievirus B3 DAF, decay accelerating factor CAR, coxsackievirus and adenovirus receptor 3Dpo1, RNA-dependent RNA polymerase.

Structural proteins derived from viruses such as HBV have the ability to spontaneously assemble into particles called virus-like particles (VLPs).These VLPs are similar to virions and consist of highly repetitive and ordered structures, but unlike recombinant protein vaccines, VLPs can elicit both humoral and cellular responses [16]. In addition these particles are easily purified and can be produced in large quantities [16]. Several VLPs are in clinical trials and have shown to be effective. This strategy has been particularly successful for HPV, and one product has been recently approved by the FDA for prophylactic use [17]. 

The study of the proteome of the recombinant adenovirus type 5 vectors demonstrated an important apphcation of separation techniques in combination with MS methods in the drug discovery process. With completely sequenced adenovirus genome available, this approach provides a chemically well-dehned method of characterization of structural proteins of recombinant adenoviral vectors. The information of protein MWs, tryptic peptide mass mapping, and sequence tags of tryptic peptides derived from HPLC/MS resulted in the identification of 17 adenoviral proteins/polypeptides in the purified virion. The rapid and accurate identification of viral proteins from recombinant adenoviruses in this study is significant since it provides direct evidence of the maturation stage of adenoviruses, which is closely related to viral infectivity and efficacy in gene therapy. 

Vaccinia virus is a large double-stranded DNA virus that infects many types of mammalian cells and some invertebrate cells. Its replication cycle is confined to the cytoplasm and can be divided into an early phase and a late phase. The late phase is characterized by the synthesis of structural proteins and enzymes that are packaged into progeny virions. Expression of foreign genes from recombinant vaccinia viruses takes advantage of promoters that are active during the late phase in order to maximize yields of the protein of interest. Several features make the vaccinia virus system... 

Expression of viral structural proteins in a heterologous system does not always lead to formation of the desired assembly intermediate or end product. This is particularly the case when the proteins are expressed in E. coli or when individual components are expressed separately in different cells. In these cases, the proteins are usually purified and then assembled in vitro. Alternatively, assembly is possible using whole cell lysates. For example, assembly of HSV-1 capsids, which requires a minimum of four structural proteins, was observed on mixing of lysates derived from insect cells infected with different baculovirus vectors (Newcomb et al, 1994). Similarly, reovirus cores, obtained from native virions, could be... 

An interesting observation was made when VPl and the minor structural protein VP2 were coexpressed in insect cells. First, immunoprecipitation analyses demonstrated a physical association between VPl and VP2, which was not surprising given the incorporation of small amounts of this protein into native virions. Second, and more interesting, the phosphorylation pattern of VPl changed in the presence of VP2. When expressed in insect cells, VPl is phosphorylated predominantly on serine, whereas in virus-infected mouse cells there is a 2 1 phosphothreonine phosphoserine ratio. On coexpression of VP2, the nonphysiologic serine phosphorylation was reduced and phosphorylation of Thr-63, the same residue that is phosphorylated in infected mouse cells, was detected (Li et al., 1995). Thus, coexpression of the two proteins in insect cells led to proper substrate phosphorylation. 

Combining these two approaches, by determining the crystal structure of individual subcomponents and fitting these into the lower resolution maps of intact particles, may offer the best current hope for obtaining high-resolution information about virion structure. An example of this is provided by adenovirus, where the crystal structure of the hexon, formed by three copies of the 100-kDa major structural protein, has been fitted into a low-resolution capsid structure determined by electron cryomicro-scopy to provide a clearer picture of capsid organization and information about the locations of minor capsid proteins (Stewart et al, 1993). A similar strategy has been used to fit the crystal structure of a portion of the HSV-1 major capsid protein (VPS) into an intermediate-resolution map of the capsid (B. Bowman, personal communication). 

Prasad and Prevelige, Fig. 3. Summary of the architectural features of rotavirus, a prototypical member of the Reoviridae family, (a) Cutaway of the mature virion structure, showing the locations of the various structural proteins, (b) Cutaway of the transcriptionally competent double-layered particle, showing the RNA core. (c). Structure of actively transcribing rotavirus double-layered particles, (d) Close-up of the exit pathway of the mRNA during transcription (Lawton et at, 1997). (e) Model for the organization of the RNA segments (Pesavento et at, 2001). 

The assembly of virions is not fully understood. It is thought that cellular factors such as chaperone proteins and the interactions between the viral nucleic acid and structural proteins might play an important role during the maturation process. 

AAV virions are small nonenveloped particles (20-25 nm) that carry a linear single-stranded DNA (ssDNA) genome, which is 4.7 kb in size. Two open reading frames (ORFs), rep and cap, have been identified in the viral genome and are flanked by T-shaped inverted terminal repeats (ITRs). The cap ORF encodes for the structural proteins that form the capsid, whereas the regulatoiy proteins are produced from the rep ORF (Figure L4). [For more details see (1).]... 

---