scholarly journals Species-Specific Conservation of Linear Antigenic Sites on Vaccinia Virus A27 Protein Homologs of Orthopoxviruses

Viruses ◽  
2019 ◽  
Vol 11 (6) ◽  
pp. 493
Author(s):  
Henrike Ahsendorf ◽  
Li Gan ◽  
Kamal Eltom ◽  
Ahmed Abd El Wahed ◽  
Sven-Kevin Hotop ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Binding Sites ◽  
Sequence Variability ◽  
Peptide Arrays ◽  
Viral Neutralization ◽  
Phylogenetic Studies ◽  
Antigenic Sites ◽  
Species Specific ◽  
Protective Proteins ◽  
Specific Conservation

The vaccinia virus (VACV) A27 protein and its homologs, which are found in a large number of members of the genus Orthopoxvirus (OPXV), are targets of viral neutralization by host antibodies. We have mapped six binding sites (epitopes #1A: aa 32–39, #1B: aa 28–33, #1C: aa 26–31, #1D: 28–34, #4: aa 9–14, and #5: aa 68–71) of A27 specific monoclonal antibodies (mAbs) using peptide arrays. MAbs recognizing epitopes #1A–D and #4 neutralized VACV Elstree in a complement dependent way (50% plaque-reduction: 12.5–200 µg/mL). Fusion of VACV at low pH was blocked through inhibition of epitope #1A. To determine the sequence variability of the six antigenic sites, 391 sequences of A27 protein homologs available were compared. Epitopes #4 and #5 were conserved among most of the OPXVs, while the sequential epitope complex #1A–D was more variable and, therefore, responsible for species-specific epitope characteristics. The accurate and reliable mapping of defined epitopes on immuno-protective proteins such as the A27 of VACV enables phylogenetic studies and insights into OPXV evolution as well as to pave the way to the development of safer vaccines and chemical or biological antivirals.

scholarly journals Structural analysis of the SRP Alu domain from Plasmodium falciparum reveals a non-canonical open conformation

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Komal Soni ◽  
Georg Kempf ◽  
Karen Manalastas-Cantos ◽  
Astrid Hendricks ◽  
Dirk Flemming ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Binding Sites ◽  
Signal Recognition Particle ◽  
Signal Recognition ◽  
Closed Conformation ◽  
Open Conformation ◽  
Structural Database ◽  
Alu Rna ◽  
Species Specific ◽  
Protozoan Species

AbstractThe eukaryotic signal recognition particle (SRP) contains an Alu domain, which docks into the factor binding site of translating ribosomes and confers translation retardation. The canonical Alu domain consists of the SRP9/14 protein heterodimer and a tRNA-like folded Alu RNA that adopts a strictly ‘closed’ conformation involving a loop-loop pseudoknot. Here, we study the structure of the Alu domain from Plasmodium falciparum (PfAlu), a divergent apicomplexan protozoan that causes human malaria. Using NMR, SAXS and cryo-EM analyses, we show that, in contrast to its prokaryotic and eukaryotic counterparts, the PfAlu domain adopts an ‘open’ Y-shaped conformation. We show that cytoplasmic P. falciparum ribosomes are non-discriminative and recognize both the open PfAlu and closed human Alu domains with nanomolar affinity. In contrast, human ribosomes do not provide high affinity binding sites for either of the Alu domains. Our analyses extend the structural database of Alu domains to the protozoan species and reveal species-specific differences in the recognition of SRP Alu domains by ribosomes.

scholarly journals Fibrinolysis: A Primordial System Linked to the Immune Response

2021 ◽  
Vol 22 (7) ◽  
pp. 3406
Author(s):  
Robert L. Medcalf ◽  
Charithani B. Keragala
Keyword(s):  
Binding Sites ◽  
Defense Mechanisms ◽  
Immune Cell ◽  
Immune Defense ◽  
Surface Proteins ◽  
Cell Behavior ◽  
Phylogenetic Studies ◽  
Blood Clots ◽  
Lower Vertebrates ◽  
Almost All

The fibrinolytic system provides an essential means to remove fibrin deposits and blood clots. The actual protease responsible for this is plasmin, formed from its precursor, plasminogen. Fibrin is heralded as it most renowned substrate but for many years plasmin has been known to cleave many other substrates, and to also activate other proteolytic systems. Recent clinical studies have shown that the promotion of plasmin can lead to an immunosuppressed phenotype, in part via its ability to modulate cytokine expression. Almost all immune cells harbor at least one of a dozen plasminogen receptors that allows plasmin formation on the cell surface that in turn modulates immune cell behavior. Similarly, a multitude of pathogens can also express their own plasminogen activators, or contain surface proteins that provide binding sites host plasminogen. Plasmin formed under these circumstances also empowers these pathogens to modulate host immune defense mechanisms. Phylogenetic studies have revealed that the plasminogen activating system predates the appearance of fibrin, indicating that plasmin did not evolve as a fibrinolytic protease but perhaps has its roots as an immune modifying protease. While its fibrin removing capacity became apparent in lower vertebrates these primitive under-appreciated immune modifying functions still remain and are now becoming more recognised.

scholarly journals Why and how might genetic and phylogenetic diversity be reflected in the identification of key biodiversity areas?

2015 ◽  
Vol 370 (1662) ◽  
pp. 20140019 ◽  
Author(s):  
T. M. Brooks ◽  
A. Cuttelod ◽  
D. P. Faith ◽  
J. Garcia-Moreno ◽  
P. Langhammer ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Threatened Species ◽  
Phylogenetic Diversity ◽  
Species Population ◽  
Threatened Taxa ◽  
Conservation Actions ◽  
Ecosystem Diversity ◽  
Species Specific ◽  
A Site ◽  
Specific Conservation

‘Key biodiversity areas' are defined as sites contributing significantly to the global persistence of biodiversity. The identification of these sites builds from existing approaches based on measures of species and ecosystem diversity and process. Here, we therefore build from the work of Sgró et al. (2011 Evol. Appl. 4 , 326–337. ( doi:10.1111/j.1752-4571.2010.00157.x )) to extend a framework for how components of genetic diversity might be considered in the identification of key biodiversity areas. We make three recommendations to inform the ongoing process of consolidating a key biodiversity areas standard: (i) thresholds for the threatened species criterion currently consider a site's share of a threatened species' population; expand these to include the proportion of the species' genetic diversity unique to a site; (ii) expand criterion for ‘threatened species' to consider ‘threatened taxa’ and (iii) expand the centre of endemism criterion to identify as key biodiversity areas those sites holding a threshold proportion of the compositional or phylogenetic diversity of species (within a taxonomic group) whose restricted ranges collectively define a centre of endemism. We also recommend consideration of occurrence of EDGE species (i.e. threatened phylogenetic diversity) in key biodiversity areas to prioritize species-specific conservation actions among sites.

THE PHOSPHOLIPID-BINDING SITE OF FACTOR VIII IS LOCATED ON THE 80 kD LIGHT CHAIN

1987 ◽  
Author(s):  
G Kemball-Cook ◽  
S J A Edwards ◽  
K Sewerin ◽  
L-O Andersson ◽  
T W Barrowcliffe
Keyword(s):  
Binding Site ◽  
Light Chain ◽  
Binding Sites ◽  
Immunoaffinity Chromatography ◽  
The Other ◽  
Immunological Methods ◽  
Electrophoretic Separation ◽  
Antigenic Sites ◽  
Reducing Conditions ◽  
Sds Page

The binding of Factoi. VIII (F.VIII) peptides to phospholipid (PL) vesicles has been studied by two different methods involving the use of fractionated anti-F.VIII:C I-Fab123’pre viously reported, i-Fab123’ was fractionated by immunoadsorptionwith F.VIII-PL complexes into two pools:one binding only to PL-binding sites on F.VIIIsAg (PL-site antibody), the other directed against other antigenic sites (non-PL-site antibody).The first technique used was a modification of the method of Weinstein et al. (Proc.Natl.Acad.Sci.USA, 78, 5137-5141, 1981), and involved incubation of the two anti-F.VIII pool swith F.VIII-containing samples, followed by electrophoretic separation of the complexes on the basis of size in non-denaturing SDS gels: this technique allows qualitative analysis of antibody reactive peptides in highly impure samples. Non-PL-site pool reacted with a range of peptides with MrMapparent Mr 90 kD up to 280 kD, a similar pattern to that of ’heavy chain’(HC) peptides of F.VIII seen on SDS-PAGE under reducing conditions; the PL-site antibody, however, reacted only with peptides at apparent Mrs of 80 kD and sometimes150 kD, but not with bands of higher Mr a pattern more consistent with binding to light chain (LC) peptides. Thesame patterns with the two labels were seen in both plasma and F.VIII concentrateThe second approach employed the two labels described above in direct immunoradiometric assays (IFMA’s) on purified human F.VIII peptides prepared by immunoaffinity chromatography and ion exchange on Mono Q gel. Both PL-site and non-PL-site labels measured similar amounts of F.VIII m a sample containing both HC and LC peptides; however, on assaying a sample containing purified HC peptides alone, PL-site antibody measured only 2% of F.VIII:Ag found by non-PL-site label, indicating that PL-binding sites present in samples containing both HC and LC are absent in HC alone.Results from both these immunological methods indicate that the 80 kD LC peptide of F.VIII carries the PL-binding site.

scholarly journals Localization of human pituitary hormones by multiple immunoenzyme staining procedures using monoclonal and polyclonal antibodies.

1986 ◽  
Vol 34 (3) ◽  
pp. 287-292 ◽  
Author(s):  
S Van Noorden ◽  
M C Stuart ◽  
A Cheung ◽  
E F Adams ◽  
J M Polak
Keyword(s):  
Alkaline Phosphatase ◽  
Binding Sites ◽  
Polyclonal Antibodies ◽  
Single Cells ◽  
Pituitary Hormones ◽  
Tissue Binding ◽  
Human Pituitary ◽  
Single Section ◽  
End Products ◽  
Species Specific

Mouse monoclonal and rabbit polyclonal antibodies to human pituitary hormones were applied together to sections of normal and neoplastic human pituitary tissue. Binding sites were revealed with species-specific immune reagents combined with various enzymes (peroxidase, alkaline phosphatase, and beta-D-galactosidase). The enzymes were developed separately to give differently colored end-products. Where two hormones were present in the same cell, a mixed color was produced. Up to four hormones could be immunostained in a single section. Multiple immunoenzymatic staining has great potential for the analysis of plural antigen production by single cells and relationships between cells producing different antigens.

scholarly journals The Relative Binding Position of Nck and Grb2 Adaptors Dramatically Impacts Actin-Based Motility of Vaccinia Virus

2021 ◽  
Author(s):  
Angika Basant ◽  
Michael Way
Keyword(s):  
Vaccinia Virus ◽  
Binding Sites ◽  
Actin Polymerization ◽  
Cellular Processes ◽  
Viral Spread ◽  
Relative Binding ◽  
Signal Output ◽  
Signalling Cascade ◽  
Binding Position

ABSTRACTPhosphotyrosine (pTyr) motifs in unstructured polypeptides orchestrate important cellular processes by engaging SH2-containing adaptors to nucleate complex signalling networks. The concept of phase separation has recently changed our appreciation of such multivalent networks, however, the role of pTyr motif positioning in their function remains to be explored. We have now explored this parameter in the assembly and operation of the signalling cascade driving actin-based motility and spread of Vaccinia virus. This network involves two pTyr motifs in the viral protein A36 that recruit the adaptors Nck and Grb2 upstream of N-WASP and Arp2/3-mediated actin polymerization. We generated synthetic networks on Vaccinia by manipulating pTyr motifs in A36 and the unrelated p14 from Orthoreovirus. In contrast to predictions, we find that only specific spatial arrangements of Grb2 and Nck binding sites result in robust N-WASP recruitment, Arp2/3 driven actin polymerization and viral spread. Our results suggest that the relative position of pTyr adaptor binding sites is optimised for signal output. This finding may explain why the relative positions of pTyr motifs are usually conserved in proteins from widely different species. It also has important implications for regulation of physiological networks, including those that undergo phase transitions.

scholarly journals Orthopoxvirus K3 orthologs show virus- and host-specific inhibition of the antiviral protein kinase PKR

2021 ◽  
Vol 17 (1) ◽  
pp. e1009183
Author(s):  
Chorong Park ◽  
Chen Peng ◽  
M. Julhasur Rahman ◽  
Sherry L. Haller ◽  
Loubna Tazi ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Vaccinia Virus ◽  
Host Range ◽  
Translation Initiation Factor ◽  
Host Cells ◽  
Initiation Factor ◽  
Specific Inhibition ◽  
Antiviral Protein ◽  
Range Function ◽  
Species Specific

The antiviral protein kinase R (PKR) is an important host restriction factor, which poxviruses must overcome to productively infect host cells. To inhibit PKR, many poxviruses encode a pseudosubstrate mimic of the alpha subunit of eukaryotic translation initiation factor 2 (eIF2), designated K3 in vaccinia virus. Although the interaction between PKR and eIF2α is highly conserved, some K3 orthologs from host-restricted poxviruses were previously shown to inhibit PKR in a species-specific manner. To better define this host range function, we compared the sensitivity of PKR from 17 mammals to inhibition by K3 orthologs from closely related orthopoxviruses, a genus with a generally broader host range. The K3 orthologs showed species-specific inhibition of PKR and exhibited three distinct inhibition profiles. In some cases, PKR from closely related species showed dramatic differences in their sensitivity to K3 orthologs. Vaccinia virus expressing the camelpox virus K3 ortholog replicated more than three orders of magnitude better in human and sheep cells than a virus expressing vaccinia virus K3, but both viruses replicated comparably well in cow cells. Strikingly, in site-directed mutagenesis experiments between the variola virus and camelpox virus K3 orthologs, we found that different amino acid combinations were necessary to mediate improved or diminished inhibition of PKR derived from different host species. Because there is likely a limited number of possible variations in PKR that affect K3-interactions but still maintain PKR/eIF2α interactions, it is possible that by chance PKR from some potential new hosts may be susceptible to K3-mediated inhibition from a virus it has never previously encountered. We conclude that neither the sensitivity of host proteins to virus inhibition nor the effectiveness of viral immune antagonists can be inferred from their phylogenetic relatedness but must be experimentally determined.

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