Recombinant Soluble Respiratory Syncytial Virus F Protein That Lacks Heptad Repeat B, Contains a GCN4 Trimerization Motif and Is Not Cleaved Displays Prefusion-Like Characteristics
The respiratory syncytial virus (RSV) fusion protein F is considered an attractive vaccine candidate especially in its prefusion conformation. We studied whether recombinant soluble RSV F proteins could be stabilized in a prefusion-like conformation by mutation of heptad repeat B (HRB). The results show that soluble, trimeric, non-cleaved RSV F protein, produced by expression of the furin cleavage site-mutated F ectodomain extended with a GCN4 trimerization sequence, is efficiently recognized by pre- as well as postfusion-specific antibodies. In contrast, a similar F protein completely lacking HRB displayed high reactivity with prefusion-specific antibodies recognizing antigenic site Ø, but did not expose postfusion-specific antigenic site I, in agreement with this protein maintaining a prefusion-like conformation. These features were dependent on the presence of the GCN4 trimerization domain. Absence of cleavage also contributed to binding of prefusion-specific antibodies. Similar antibody reactivity profiles were observed when the prefusion form of F was stabilized by the introduction of cysteine pairs in HRB. To study whether the inability to form the 6HB was responsible for the prefusion-like antibody reactivity profile, alanine mutations were introduced in HRB. Although introduction of alanine residues in HRB inhibited the formation of the 6HB, the exposure of postfusion-specific antigenic site I was not prevented. In conclusion, proteins that are not able to form the 6HB, due to mutation of HRB, may still display postfusion-specific antigenic site I. Replacement of HRB by the GCN4 trimerization domain in a non-cleaved soluble F protein resulted, however, in a protein with prefusion-like characteristics, suggesting that this HRB-lacking protein may represent a potential prefusion F protein subunit vaccine candidate.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2015 |
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| Erschienen: |
2015 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:10 |
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| Enthalten in: |
PloS one - 10(2015), 6 vom: 28., Seite e0130829 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Widjaja, Ivy [VerfasserIn] |
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| Links: |
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| Themen: |
Antibodies, Neutralizing |
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| Anmerkungen: |
Date Completed 23.03.2016 Date Revised 13.11.2018 published: Electronic-eCollection Citation Status MEDLINE |
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| doi: |
10.1371/journal.pone.0130829 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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NLM250273535 |
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| 245 | 1 | 0 | |a Recombinant Soluble Respiratory Syncytial Virus F Protein That Lacks Heptad Repeat B, Contains a GCN4 Trimerization Motif and Is Not Cleaved Displays Prefusion-Like Characteristics |
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| 500 | |a Citation Status MEDLINE | ||
| 520 | |a The respiratory syncytial virus (RSV) fusion protein F is considered an attractive vaccine candidate especially in its prefusion conformation. We studied whether recombinant soluble RSV F proteins could be stabilized in a prefusion-like conformation by mutation of heptad repeat B (HRB). The results show that soluble, trimeric, non-cleaved RSV F protein, produced by expression of the furin cleavage site-mutated F ectodomain extended with a GCN4 trimerization sequence, is efficiently recognized by pre- as well as postfusion-specific antibodies. In contrast, a similar F protein completely lacking HRB displayed high reactivity with prefusion-specific antibodies recognizing antigenic site Ø, but did not expose postfusion-specific antigenic site I, in agreement with this protein maintaining a prefusion-like conformation. These features were dependent on the presence of the GCN4 trimerization domain. Absence of cleavage also contributed to binding of prefusion-specific antibodies. Similar antibody reactivity profiles were observed when the prefusion form of F was stabilized by the introduction of cysteine pairs in HRB. To study whether the inability to form the 6HB was responsible for the prefusion-like antibody reactivity profile, alanine mutations were introduced in HRB. Although introduction of alanine residues in HRB inhibited the formation of the 6HB, the exposure of postfusion-specific antigenic site I was not prevented. In conclusion, proteins that are not able to form the 6HB, due to mutation of HRB, may still display postfusion-specific antigenic site I. Replacement of HRB by the GCN4 trimerization domain in a non-cleaved soluble F protein resulted, however, in a protein with prefusion-like characteristics, suggesting that this HRB-lacking protein may represent a potential prefusion F protein subunit vaccine candidate | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 7 | |a Antibodies, Neutralizing |2 NLM | |
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| 700 | 1 | |a Rigter, Alan |e verfasserin |4 aut | |
| 700 | 1 | |a Jacobino, Shamir |e verfasserin |4 aut | |
| 700 | 1 | |a van Kuppeveld, Frank J M |e verfasserin |4 aut | |
| 700 | 1 | |a Leenhouts, Kees |e verfasserin |4 aut | |
| 700 | 1 | |a Palomo, Concepción |e verfasserin |4 aut | |
| 700 | 1 | |a Melero, Jose A |e verfasserin |4 aut | |
| 700 | 1 | |a Leusen, Jeanette H W |e verfasserin |4 aut | |
| 700 | 1 | |a Haijema, Bert Jan |e verfasserin |4 aut | |
| 700 | 1 | |a Rottier, Peter J M |e verfasserin |4 aut | |
| 700 | 1 | |a de Haan, Cornelis A M |e verfasserin |4 aut | |
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