Einfluss gezielter Mutationen auf die biologische Aktivität des Oberflächen-Glykoproteins eines afrikanischen Henipavirus
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Philipps-Universität Marburg
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Abstract
Hendra virus and Nipah virus are the two highly pathogenic representatives of the genus
henipavirus. Fruit bats of the genus Pteropus serve as their natural reservoir. While bats
do not show any clinical symptoms upon infection, henipaviruses can cause severe
illness after natural spillover to humans, pigs or horses. Due to their zoonotic potential
and high pathogenicity, henipaviruses are classified as BSL-4 pathogens. Initially,
henipaviruses were believed to be restricted to Southeast Asia and Australia. But in
recent years, there is increasing evidence of a much broaden geographic distribution
because henipaviral RNA has been identified also in Africa. One of these African bat
henipaviruses, Kumasi virus (KV), isolated from an Eidolon helvum fruit bat in Ghana,
could be fully sequenced. Since isolation of infectious viruses was not successful so far,
evaluation of the zoonotic potential of new viruses is based on functional analyses of
individual viral surface proteins in comparison to their pathogenic homologues from
Nipah or Hendra viruses.
The receptor-binding G protein and the fusion protein F are central determinants for virus
entry and cell-to-cell spread. Efficient binding to cellular receptors and fusion-helper
function of henipavirus G proteins are essentially required to support F-mediated viruscell and cell-cell-fusion processes. It has been shown earlier that surface expression and
fusion-helper activity of KV-G is clearly reduced compared to the G protein of pathogenic
NiV (NiV-G). To gain insight into the molecular determinants affecting protein transport
and function, several KV-G mutants resulting in changes in N glycosylation,
oligomerization and endocytosis pattern were generated. Western Blot analysis,
metabolic labeling and functional fusion assays revealed that, similar to NiV-G, KV-G
contains six N glycans which are all required for surface transport and functionality.
However, KV-G differs in its oligomerization by almost exclusively forming stable
tetramers, while NiV-G is expressed in a well-balanced dimer-tetramer ratio. Although
cysteine mutations in the stalk domain of KV-G altered the oligomerization pattern,
neither surface transport nor fusion-helper activity was improved. However, when
mutating a non-conserved cysteine in the globular head domain of KV-G some gain of
function was observed. This was further increased when we additionally disrupted an
endocytosis motive in the cytoplasmic domain. This work shows for the first time an
African henipavirus glycoprotein with enhanced functionality. Although the bioactivity is
still far below that of highly pathogenic henipavirus G proteins, this indicates that African
henipaviruses with improved fusion-helper function and perhaps an increased zoonotic
potential might evolve by just a few adaptive mutations.
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Created: 2017Issued: 2018-10-15Updated: 2018-10-15
Faculty
Fachbereich Biologie
Publisher
Philipps-Universität Marburg
Language
ger
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DoctoralThesis
Keywords
Fusionshelferfunktionsurface expressionOligomerisierungoligomerizationOberflächenexpressionHenipavirusglycoproteinfusion-helper functionHenipavirusGlykoprotein
DFG-subjects
GlykoproteinOligomerisierungOberflächenexpressionHenipavirusFusionshelferfunktion
DDC-Numbers
570
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Behner, Laura: Einfluss gezielter Mutationen auf die biologische Aktivität des Oberflächen-Glykoproteins eines afrikanischen Henipavirus. : Philipps-Universität Marburg 2018-10-15. DOI: https://doi.org/10.17192/z2018.0097.
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This item has been published with the following license: In Copyright