scholarly journals Structural analysis of the human respiratory syncytial virus phosphoprotein: characterization of an α-helical domain involved in oligomerization

2006 ◽  
Vol 87 (1) ◽  
pp. 159-169 ◽  
Author(s):  
María T. Llorente ◽  
Blanca García-Barreno ◽  
Miguel Calero ◽  
Emilio Camafeita ◽  
Juan A. López ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Mammalian Cells ◽  
Gel Filtration ◽  
Human Respiratory Syncytial Virus ◽  
Mass Spectrometry Analysis ◽  
Modular Organization ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Cell Extracts ◽  
Syncytial Virus

Human respiratory syncytial virus (HRSV) phosphoprotein (P), an essential cofactor of the viral polymerase, is much shorter (241 aa) than and has no sequence similarity to P of other paramyxoviruses. Nevertheless, bioinformatic analysis of HRSV P sequence revealed a modular organization, reminiscent of other paramyxovirus Ps, with a central structured domain (aa 100–200), flanked by two intrinsically disordered regions (1–99 and 201–241). To test the predicted structure experimentally, HRSV P was purified from cell extracts infected with recombinant vaccinia virus or HRSV. The estimated molecular mass of P by gel filtration (∼500 kDa) greatly exceeded the theoretical mass of a homotetramer, proposed as the oligomeric form of native P. Nevertheless, the profile of cross-linked products obtained with purified P resembled that reported by others with P purified from bacteria or mammalian cells. Thus, the shape of HRSV P probably influences its elution from the gel filtration column, as reported for other paramyxovirus Ps. Digestion of purified HRSV P with different proteases identified a trypsin-resistant fragment (X) that reacted with a previously characterized monoclonal antibody (021/2P). N-terminal sequencing and mass spectrometry analysis placed the X fragment boundaries (Glu-104 and Arg-163) within the predicted structured domain of P. Cross-linking and circular dichroism analyses indicated that fragment X was oligomeric, with a high α-helical content, properties resembling those of the multimerization domain of Sendai and rinderpest virus P. These results denote structural features shared by HRSV and other paramyxovirus Ps and should assist in elucidation of the HRSV P structure.

scholarly journals The Baculovirus GP64 Protein Mediates Highly Stable Infectivity of a Human Respiratory Syncytial Virus Lacking Its Homologous Transmembrane Glycoproteins

2004 ◽  
Vol 78 (1) ◽  
pp. 124-135 ◽  
Author(s):  
A. G. P. Oomens ◽  
Gail W. Wertz
Keyword(s):  
Respiratory Syncytial Virus ◽  
Membrane Fusion ◽  
Mammalian Cells ◽  
Chimeric Protein ◽  
Low Ph ◽  
Human Respiratory Syncytial Virus ◽  
Dependent Manner ◽  
Practical Applications ◽  
Ph Dependent ◽  
Syncytial Virus

ABSTRACT Baculovirus GP64 is a low-pH-dependent membrane fusion protein required for virus entry and cell-to-cell transmission. Recently, GP64 has generated interest for practical applications in mammalian systems. Here we examined the membrane fusion function of GP64 from Autographa californica multiple nucleopolyhedrovirus (AcMNPV) expressed in mammalian cells, as well as its capacity to functionally complement a mammalian virus, human respiratory syncytial virus (HRSV). Both authentic GP64 and GP64/F, a chimeric protein in which the GP64 cytoplasmic tail domain was replaced with the 12 C-terminal amino acids of the HRSV fusion (F) protein, induced low-pH-dependent cell-cell fusion when expressed transiently in HEp-2 (human) cells. Levels of surface expression and syncytium formation were substantially higher at 33°C than at 37°C. The open reading frames (ORFs) encoding GP64 or GP64/F, along with two marker ORFs encoding green fluorescent protein (GFP) and β-glucuronidase (GUS), were used to replace all three homologous transmembrane glycoprotein ORFs (small hydrophobic SH, attachment G, and F) in a cDNA of HRSV. Infectious viruses were recovered that lacked the HRSV SH, G, and F proteins and expressed instead the GP64 or GP64/F protein and the two marker proteins GFP and GUS. The properties of these viruses, designated RSΔsh,g,f/GP64 or RSΔsh,g,f/GP64/F, respectively, were compared to a previously described HRSV expressing GFP in place of SH but still containing the wild-type HRSV G and F proteins (RSΔsh [A. G. Oomens, A. G. Megaw, and G. W. Wertz, J. Virol., 77:3785-3798, 2003]). By immunoelectron microscopy, the GP64 and GP64/F proteins were shown to incorporate into HRSV-induced filaments at the cell surface. Antibody neutralization, ammonium chloride inhibition, and replication levels in cell culture showed that both GP64 proteins efficiently mediated infectivity of the respective viruses in a temperature-sensitive, low-pH-dependent manner. Furthermore, RSΔsh,g,f/GP64 and RSΔsh,g,f/GP64/F replicated to higher levels and had significantly higher stability of infectivity than HRSVs containing the homologous HRSV G and F proteins. Thus, GP64 and a GP64/HRSV F chimeric protein were functional and efficiently complemented an unrelated human virus in mammalian cells, producing stable, infectious virus stocks. These results demonstrate the potential of GP64 for both practical applications requiring stable pseudotypes in mammalian systems and for studies of viral glycoprotein requirements in assembly and pathogenesis.

scholarly journals Functional correlations of respiratory syncytial virus proteins to intrinsic disorder

2016 ◽  
Vol 12 (5) ◽  
pp. 1507-1526 ◽  
Author(s):  
Jillian N. Whelan ◽  
Krishna D. Reddy ◽  
Vladimir N. Uversky ◽  
Michael N. Teng
Keyword(s):  
Respiratory Syncytial Virus ◽  
Intrinsic Disorder ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Syncytial Virus ◽  
Disordered Regions ◽  
Virus Proteins

The respiratory syncytial virus proteome is highly enriched in intrinsically disordered regions, which confer many functional advantages.

scholarly journals Evaluation of the Safety and Immune Efficacy of Recombinant Human Respiratory Syncytial Virus Strain Long Live Attenuated Vaccine Candidates

2021 ◽  
Author(s):  
Li-Nan Wang ◽  
Xiang-Lei Peng ◽  
Min Xu ◽  
Yuan-Bo Zheng ◽  
Yue-Ying Jiao ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Gene Deletion ◽  
Human Respiratory Syncytial Virus ◽  
Temperature Sensitive ◽  
T7 Promoter ◽  
Vaccine Candidates ◽  
Rsv Infection ◽  
Syncytial Virus

AbstractHuman respiratory syncytial virus (RSV) infection is the leading cause of lower respiratory tract illness (LRTI), and no vaccine against LRTI has proven to be safe and effective in infants. Our study assessed attenuated recombinant RSVs as vaccine candidates to prevent RSV infection in mice. The constructed recombinant plasmids harbored (5′ to 3′) a T7 promoter, hammerhead ribozyme, RSV Long strain antigenomic cDNA with cold-passaged (cp) mutations or cp combined with temperature-sensitive attenuated mutations from the A2 strain (A2cpts) or further combined with SH gene deletion (A2cptsΔSH), HDV ribozyme (δ), and a T7 terminator. These vectors were subsequently co-transfected with four helper plasmids encoding N, P, L, and M2-1 viral proteins into BHK/T7-9 cells, and the recovered viruses were then passaged in Vero cells. The rescued recombinant RSVs (rRSVs) were named rRSV-Long/A2cp, rRSV-Long/A2cpts, and rRSV-Long/A2cptsΔSH, respectively, and stably passaged in vitro, without reversion to wild type (wt) at sites containing introduced mutations or deletion. Although rRSV-Long/A2cpts and rRSV-Long/A2cptsΔSH displayed  temperature-sensitive (ts) phenotype in vitro and in vivo, all rRSVs were significantly attenuated in vivo. Furthermore, BALB/c mice immunized with rRSVs produced Th1-biased immune response, resisted wtRSV infection, and were free from enhanced respiratory disease. We showed that the combination of ΔSH with attenuation (att) mutations of cpts contributed to improving att phenotype, efficacy, and gene stability of rRSV. By successfully introducing att mutations and SH gene deletion into the RSV Long parent and producing three rRSV strains, we have laid an important foundation for the development of RSV live attenuated vaccines.

scholarly journals Fatty Acid Acylation of the Fusion Glycoprotein of Human Respiratory Syncytial Virus

1989 ◽  
Vol 264 (18) ◽  
pp. 10339-10342
Author(s):  
R G Arumugham ◽  
R C Seid ◽  
S Doyle ◽  
S W Hildreth ◽  
P R Paradiso
Keyword(s):  
Fatty Acid ◽  
Respiratory Syncytial Virus ◽  
Human Respiratory Syncytial Virus ◽  
Fusion Glycoprotein ◽  
Syncytial Virus

scholarly journals Bacterial and Viral Coinfections with the Human Respiratory Syncytial Virus

2021 ◽  
Vol 9 (6) ◽  
pp. 1293
Author(s):  
Gaspar A. Pacheco ◽  
Nicolás M. S. Gálvez ◽  
Jorge A. Soto ◽  
Catalina A. Andrade ◽  
Alexis M. Kalergis
Keyword(s):  
Respiratory Syncytial Virus ◽  
Influenza A ◽  
Human Respiratory Syncytial Virus ◽  
Respiratory Pathogens ◽  
Parainfluenza Viruses ◽  
Starting Point ◽  
The Social ◽  
Syncytial Virus ◽  
Tract Infections ◽  
Polymicrobial Infections

The human respiratory syncytial virus (hRSV) is one of the leading causes of acute lower respiratory tract infections in children under five years old. Notably, hRSV infections can give way to pneumonia and predispose to other respiratory complications later in life, such as asthma. Even though the social and economic burden associated with hRSV infections is tremendous, there are no approved vaccines to date to prevent the disease caused by this pathogen. Recently, coinfections and superinfections have turned into an active field of study, and interactions between many viral and bacterial pathogens have been studied. hRSV is not an exception since polymicrobial infections involving this virus are common, especially when illness has evolved into pneumonia. Here, we review the epidemiology and recent findings regarding the main polymicrobial infections involving hRSV and several prevalent bacterial and viral respiratory pathogens, such as Staphylococcus aureus, Pseudomonas aeruginosa, Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, Klebsiella pneumoniae, human rhinoviruses, influenza A virus, human metapneumovirus, and human parainfluenza viruses. As reports of most polymicrobial infections involving hRSV lack a molecular basis explaining the interaction between hRSV and these pathogens, we believe this review article can serve as a starting point to interesting and very much needed research in this area.

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