scholarly journals Roles of the highly conserved amino acids in the globular head and stalk region of the Newcastle disease virus HN protein in the membrane fusion process

2015 ◽  
Vol 9 (1) ◽  
pp. 56-64 ◽  
Author(s):  
Chengxi Sun ◽  
Hongling Wen ◽  
Yuzhen Chen ◽  
Fulu Chu ◽  
Bin Lin ◽  
...  
Keyword(s):  
Amino Acids ◽  
Newcastle Disease Virus ◽  
Membrane Fusion ◽  
Disease Virus ◽  
Newcastle Disease ◽  
Fusion Process ◽  
Conserved Amino Acids ◽  
Globular Head

scholarly journals Roles of the highly conserved amino acids in the second receptor binding site of the Newcastle disease virus HN protein

2019 ◽  
Vol 16 (1) ◽  
Author(s):  
Yaqing Liu ◽  
Miaomiao Chi ◽  
Ying Liu ◽  
Hongling Wen ◽  
Li Zhao ◽  
...  
Keyword(s):  
Amino Acids ◽  
Newcastle Disease Virus ◽  
Binding Site ◽  
Conformational Change ◽  
Disease Virus ◽  
Receptor Binding ◽  
Newcastle Disease ◽  
Cell Surface Expression ◽  
Receptor Binding Site ◽  
Conserved Amino Acids

Abstract Background The paramyxovirus haemagglutinin-neuraminidase (HN) is a multifunctional protein that is responsible for attachment to receptors, removal of receptors from infected cells to prevent viral self-aggregation (neuraminidase, NA) and fusion promotion. It is commonly accepted that there are two receptor binding sites in the globular head of HN, and the second receptor binding site is only involved in the function of receptor binding and fusion promotion. Methods 10 conserved residues in the second receptor binding site of Newcastle disease virus (NDV) HN were chosen and substituted to alanine (A). The desired mutants were examined to detect the functional change in hemadsorption (HAD) ability, NA activity and fusion promotion ability. Results The HAD and fusion promotion ability of mutants C172A, R174A, C196A, D198A, Y526A and E547A were abolished. Compared with wild-type (wt) HN, the HAD of mutants T167A, S202A and R516A decreased to 55.81, 44.53, 69.02%, respectively, and the fusion promotion ability of these three mutants decreased to 54.74, 49.46, 65.26%, respectively; however, mutant G171A still maintained fusion promotion ability comparable with wt HN but had impaired HAD ability. All the site-directed mutations altered the NA activity of NDV HN without affecting protein cell surface expression. Conclusions The data suggest that mutants C172A, R174A, C196A, D198A, Y526A and E547A do not allow the conformational change that is required for fusion promotion ability and HAD activity, while the other mutants only affect the conformational change to a limited extent, except mutant G171A with intact fusion promotion ability. Overall, the conserved amino acids in the second receptor binding site, especially residues C172, R174, C196, D198, Y526 and E547, are crucial to normal NDV HN protein function.

scholarly journals Addition of N-Glycans in the Stalk of the Newcastle Disease Virus HN Protein Blocks Its Interaction with the F Protein and Prevents Fusion

2006 ◽  
Vol 80 (2) ◽  
pp. 623-633 ◽  
Author(s):  
Vanessa R. Melanson ◽  
Ronald M. Iorio
Keyword(s):  
Newcastle Disease Virus ◽  
Membrane Fusion ◽  
Newcastle Disease ◽  
Sucrose Gradient ◽  
Specific Interaction ◽  
F Protein ◽  
Tetrameric Structure ◽  
Protein Blocks ◽  
Globular Head

ABSTRACT Most paramyxovirus fusion (F) proteins require the coexpression of the homologous attachment (HN) protein to promote membrane fusion, consistent with the existence of a virus-specific interaction between the two proteins. Analysis of the fusion activities of chimeric HN proteins indicates that the stalk region of the HN spike determines its F protein specificity, and analysis of a panel of site-directed mutants indicates that the F-interactive site resides in this region. Here, we use the addition of oligosaccharides to further explore the role of the HN stalk in the interaction with F. N-glycans were individually added at several positions in the stalk to determine their effects on the activities of HN, as well as its structure. N-glycan addition at positions 69 and 77 in the stalk specifically blocks fusion and the HN-F interaction without affecting either HN structure or its other activities. N-glycans added at other positions in the stalk modulate activities that reside in the globular head of HN. This correlates with an alteration of the tetrameric structure of the protein, as indicated by sucrose gradient sedimentation analyses. Finally, N-glycan addition in another region of HN (residues 124 to 152), predicted by a peptide-based analysis to mediate the interaction with F, does not significantly reduce the level of fusion, arguing strongly against this site being part of the F-interactive domain in HN. Our data support the idea that the F-interactive site on HN is defined by the stalk region of the protein.

scholarly journals Effect of Cleavage Mutants on Syncytium Formation Directed by the Wild-Type Fusion Protein of Newcastle Disease Virus

1998 ◽  
Vol 72 (5) ◽  
pp. 3789-3795 ◽  
Author(s):  
Zengji Li ◽  
Theresa Sergel ◽  
Enal Razvi ◽  
Trudy Morrison
Keyword(s):  
Amino Acids ◽  
Newcastle Disease Virus ◽  
Disease Virus ◽  
Newcastle Disease ◽  
Syncytium Formation ◽  
Dominant Negative ◽  
Wild Type ◽  
F Protein ◽  
Type Protein ◽  
Wild Type Protein

ABSTRACT The effects of Newcastle disease virus (NDV) fusion (F) glycoprotein cleavage mutants on the cleavage and syncytium-forming activity of the wild-type F protein were examined. F protein cleavage mutants were made by altering amino acids in the furin recognition region (amino acids 112 to 116) in the F protein of a virulent strain of NDV. Four mutants were made: Q114P replaced the glutamine residue with proline; K115G replaced lysine with glycine; double mutant K115G, R113G replaced both a lysine and an arginine with glycine residues; and a triple mutant, R112G, K115G, F117L, replaced three amino acids to mimic the sequence found in avirulent strains of NDV. All mutants except Q114P were cleavage negative and fusion negative. However, addition of exogenous trypsin cleaved all mutant F proteins and activated fusion. As expected for an oligomeric protein, the fusion-negative mutants had a dominant negative phenotype: cotransfection of wild-type and mutant F protein cDNAs resulted in an inhibition of syncytium formation. The presence of the mutant F protein did not inhibit cleavage of the wild-type protein. Furthermore, evidence is presented that suggests that the mutant protein and the wild-type protein formed heterooligomers. By measuring the syncytium-forming activity of the wild-type protein at various ratios of expression of mutant and wild-type protein, results were obtained that are most consistent with the notion that the size of the functionally active NDV F protein in these assays is a single oligomer, likely a trimer. That a larger oligomer, containing a mix of both wild-type and mutant F proteins, has partial activity cannot, however, be ruled out.

scholarly journals Virulence of Newcastle disease virus is determined by the cleavage site of the fusion protein and by both the stem region and globular head of the haemagglutinin–neuraminidase protein

2005 ◽  
Vol 86 (6) ◽  
pp. 1759-1769 ◽  
Author(s):  
Olav S. de Leeuw ◽  
Guus Koch ◽  
Leo Hartog ◽  
Niek Ravenshorst ◽  
Ben P. H. Peeters
Keyword(s):  
Newcastle Disease Virus ◽  
Disease Virus ◽  
Newcastle Disease ◽  
Cleavage Site ◽  
Infectious Clone ◽  
Protein Cleavage ◽  
F Protein ◽  
Virulent Strains ◽  
Intravenous Inoculation ◽  
Globular Head

Virulence of Newcastle disease virus (NDV) is mainly determined by the amino acid sequence surrounding the fusion (F) protein cleavage site, since host proteases that cleave the F protein of virulent strains are present in more tissues than those that cleave the F protein of non-virulent strains. Nevertheless, comparison of NDV strains that carry exactly the same F protein cleavage site shows that significant differences in virulence still exist. For instance, virulent field strain Herts/33 with the F cleavage site 112RRQRRF117 had an intracerebral pathogenicity index of 1·88 compared with 1·28 for strain NDFLtag, which has the same cleavage site. This implies that additional factors contribute to virulence. After generating an infectious clone of Herts/33 (FL-Herts), we were able to map the location of additional virulence factors by exchanging sequences between FL-Herts and NDFLtag. The results showed that, in addition to the F protein cleavage site, the haemagglutinin–neuraminidase (HN) protein also contributed to virulence. The effect of the HN protein on virulence was most prominent after intravenous inoculation. Interestingly, both the stem region and the globular head of the HN protein seem to be involved in determining virulence.

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