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Identification of specific histidines as pH sensors in flavivirus membrane fusion.

Fritz R, Stiasny K, Heinz FX - J. Cell Biol. (2008)

Bottom Line: It has been hypothesized that conserved histidines in the class II fusion protein E of these viruses function as molecular switches and, by their protonation, control the fusion process.Using the mutational analysis of recombinant subviral particles of tick-borne encephalitis virus, we provide direct experimental evidence that the initiation of fusion is crucially dependent on the protonation of one of the conserved histidines (His323) at the interface between domains I and III of E, leading to the dissolution of domain interactions and to the exposure of the fusion peptide.Conserved histidines located outside this critical interface were found to be completely dispensable for triggering fusion.

View Article: PubMed Central - PubMed

Affiliation: Institute of Virology, Medical University of Vienna, 1095 Vienna, Austria.

ABSTRACT
The flavivirus membrane fusion machinery, like that of many other enveloped viruses, is triggered by the acidic pH in endosomes after virus uptake by receptor-mediated endocytosis. It has been hypothesized that conserved histidines in the class II fusion protein E of these viruses function as molecular switches and, by their protonation, control the fusion process. Using the mutational analysis of recombinant subviral particles of tick-borne encephalitis virus, we provide direct experimental evidence that the initiation of fusion is crucially dependent on the protonation of one of the conserved histidines (His323) at the interface between domains I and III of E, leading to the dissolution of domain interactions and to the exposure of the fusion peptide. Conserved histidines located outside this critical interface were found to be completely dispensable for triggering fusion.

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Summary of the organization of flavivirus particles, the three-dimensional structures of the flavivirus envelope protein E, and a model of flavivirus membrane fusion. (A) Schematic diagram of a flavivirus particle in its immature (prM-containing) and mature form after proteolytic cleavage of prM. (B) Schematic of the prefusion E dimer including ribbon diagrams of the TBEV sE ectodomain (top and side view) and those parts for which the atomic structure is not known (stem and anchor). (C) Ribbon diagram of the postfusion TBEV sE trimer (side view). The positions of the histidines conserved in all flavivirus E proteins are indicated by gray balls. (D) Schematic of the proposed flavivirus fusion mechanism showing different steps of the fusion process. (step 1) Metastable E dimer in mature virions. (step 2) Dissociation of the E dimers at acidic pH, outward projection of E monomers, and interaction of the FP with the target membrane. (step 3) Trimerization, DIII relocation, and “zipping up” of the stem. (step 4) Formation of the postfusion trimer and opening of the fusion pore. Red, DI; yellow, DII; blue, DIII; orange, FP; purple, stem (linker between DIII and the transmembrane anchors); gray, transmembrane anchors.
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fig1: Summary of the organization of flavivirus particles, the three-dimensional structures of the flavivirus envelope protein E, and a model of flavivirus membrane fusion. (A) Schematic diagram of a flavivirus particle in its immature (prM-containing) and mature form after proteolytic cleavage of prM. (B) Schematic of the prefusion E dimer including ribbon diagrams of the TBEV sE ectodomain (top and side view) and those parts for which the atomic structure is not known (stem and anchor). (C) Ribbon diagram of the postfusion TBEV sE trimer (side view). The positions of the histidines conserved in all flavivirus E proteins are indicated by gray balls. (D) Schematic of the proposed flavivirus fusion mechanism showing different steps of the fusion process. (step 1) Metastable E dimer in mature virions. (step 2) Dissociation of the E dimers at acidic pH, outward projection of E monomers, and interaction of the FP with the target membrane. (step 3) Trimerization, DIII relocation, and “zipping up” of the stem. (step 4) Formation of the postfusion trimer and opening of the fusion pore. Red, DI; yellow, DII; blue, DIII; orange, FP; purple, stem (linker between DIII and the transmembrane anchors); gray, transmembrane anchors.

Mentions: Flaviviruses (genus Flavivirus and family Flaviviridae) have an acidic pH–dependent fusion machinery (Stiasny and Heinz, 2006) and comprise several closely related important human pathogens, including yellow fever, dengue, Japanese encephalitis, West Nile, and TBE viruses (Gubler et al., 2007). The surface of mature flaviviruses is made up of a herringbone-like assembly of 90 homodimers of the envelope glycoprotein E (Kuhn et al., 2002; Mukhopadhyay et al., 2003). The atomic structures of soluble forms of E (sE), lacking the membrane anchor and the so-called stem (Fig. 1 B), have been determined for different flaviviruses in pre- and postfusion conformations (Fig. 1, B and C; Rey et al., 1995; Modis et al., 2003, 2004, 2005; Bressanelli et al., 2004; Zhang et al., 2004; Kanai et al., 2006; Nybakken et al., 2006). In the prefusion conformation, the internal fusion peptide (FP) loop at the tip of domain II (DII) is buried through the interaction with a hydrophobic pocket provided by DI and III of the second partner in the homodimer (Fig. 1 B). Exposure to acidic pH leads to the initiation of the fusion process as depicted in Fig. 1 D.


Identification of specific histidines as pH sensors in flavivirus membrane fusion.

Fritz R, Stiasny K, Heinz FX - J. Cell Biol. (2008)

Summary of the organization of flavivirus particles, the three-dimensional structures of the flavivirus envelope protein E, and a model of flavivirus membrane fusion. (A) Schematic diagram of a flavivirus particle in its immature (prM-containing) and mature form after proteolytic cleavage of prM. (B) Schematic of the prefusion E dimer including ribbon diagrams of the TBEV sE ectodomain (top and side view) and those parts for which the atomic structure is not known (stem and anchor). (C) Ribbon diagram of the postfusion TBEV sE trimer (side view). The positions of the histidines conserved in all flavivirus E proteins are indicated by gray balls. (D) Schematic of the proposed flavivirus fusion mechanism showing different steps of the fusion process. (step 1) Metastable E dimer in mature virions. (step 2) Dissociation of the E dimers at acidic pH, outward projection of E monomers, and interaction of the FP with the target membrane. (step 3) Trimerization, DIII relocation, and “zipping up” of the stem. (step 4) Formation of the postfusion trimer and opening of the fusion pore. Red, DI; yellow, DII; blue, DIII; orange, FP; purple, stem (linker between DIII and the transmembrane anchors); gray, transmembrane anchors.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2568029&req=5

fig1: Summary of the organization of flavivirus particles, the three-dimensional structures of the flavivirus envelope protein E, and a model of flavivirus membrane fusion. (A) Schematic diagram of a flavivirus particle in its immature (prM-containing) and mature form after proteolytic cleavage of prM. (B) Schematic of the prefusion E dimer including ribbon diagrams of the TBEV sE ectodomain (top and side view) and those parts for which the atomic structure is not known (stem and anchor). (C) Ribbon diagram of the postfusion TBEV sE trimer (side view). The positions of the histidines conserved in all flavivirus E proteins are indicated by gray balls. (D) Schematic of the proposed flavivirus fusion mechanism showing different steps of the fusion process. (step 1) Metastable E dimer in mature virions. (step 2) Dissociation of the E dimers at acidic pH, outward projection of E monomers, and interaction of the FP with the target membrane. (step 3) Trimerization, DIII relocation, and “zipping up” of the stem. (step 4) Formation of the postfusion trimer and opening of the fusion pore. Red, DI; yellow, DII; blue, DIII; orange, FP; purple, stem (linker between DIII and the transmembrane anchors); gray, transmembrane anchors.
Mentions: Flaviviruses (genus Flavivirus and family Flaviviridae) have an acidic pH–dependent fusion machinery (Stiasny and Heinz, 2006) and comprise several closely related important human pathogens, including yellow fever, dengue, Japanese encephalitis, West Nile, and TBE viruses (Gubler et al., 2007). The surface of mature flaviviruses is made up of a herringbone-like assembly of 90 homodimers of the envelope glycoprotein E (Kuhn et al., 2002; Mukhopadhyay et al., 2003). The atomic structures of soluble forms of E (sE), lacking the membrane anchor and the so-called stem (Fig. 1 B), have been determined for different flaviviruses in pre- and postfusion conformations (Fig. 1, B and C; Rey et al., 1995; Modis et al., 2003, 2004, 2005; Bressanelli et al., 2004; Zhang et al., 2004; Kanai et al., 2006; Nybakken et al., 2006). In the prefusion conformation, the internal fusion peptide (FP) loop at the tip of domain II (DII) is buried through the interaction with a hydrophobic pocket provided by DI and III of the second partner in the homodimer (Fig. 1 B). Exposure to acidic pH leads to the initiation of the fusion process as depicted in Fig. 1 D.

Bottom Line: It has been hypothesized that conserved histidines in the class II fusion protein E of these viruses function as molecular switches and, by their protonation, control the fusion process.Using the mutational analysis of recombinant subviral particles of tick-borne encephalitis virus, we provide direct experimental evidence that the initiation of fusion is crucially dependent on the protonation of one of the conserved histidines (His323) at the interface between domains I and III of E, leading to the dissolution of domain interactions and to the exposure of the fusion peptide.Conserved histidines located outside this critical interface were found to be completely dispensable for triggering fusion.

View Article: PubMed Central - PubMed

Affiliation: Institute of Virology, Medical University of Vienna, 1095 Vienna, Austria.

ABSTRACT
The flavivirus membrane fusion machinery, like that of many other enveloped viruses, is triggered by the acidic pH in endosomes after virus uptake by receptor-mediated endocytosis. It has been hypothesized that conserved histidines in the class II fusion protein E of these viruses function as molecular switches and, by their protonation, control the fusion process. Using the mutational analysis of recombinant subviral particles of tick-borne encephalitis virus, we provide direct experimental evidence that the initiation of fusion is crucially dependent on the protonation of one of the conserved histidines (His323) at the interface between domains I and III of E, leading to the dissolution of domain interactions and to the exposure of the fusion peptide. Conserved histidines located outside this critical interface were found to be completely dispensable for triggering fusion.

Show MeSH
Related in: MedlinePlus