Page 57 - TBE_Book_V2_2019

Chapter 2b: The molecular and antigenic structure of TBEV

acidic endosomal compartment for viral mechanism consists of several steps as depicted
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membrane fusion. The acid pH-dependence of in Figure 3C. In this process, the acidic-pH-
TBEV fusion activity was first demonstrated by induced dissociation of E dimers leads to the
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Guirakhoo et al. 1991 and further studied in exposure and interaction of the FL with the
great detail using a combination of biochemical, endosomal membrane, the relocation of DIII
structural, mutational, and functional and the zippering of the ‘stem’ along DII in the
studies. 54,57,59,60,78-83 Chemical cross-linking trimer, thus driving the merger of the two
experiments and sedimentation analyses membranes. Mutational analyses provided
demonstrated that the exposure to acidic pH evidence for a specific molecular interaction at
caused a quantitative oligomeric rearrangement the N-terminal end of the stem and a pocket of
of metastable E dimers into stable trimers at DII which appears to be essential for the correct
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the virion surface, with a pH threshold of 6.5, positioning of the stem for the zippering
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suggesting that this dimer-trimer transition reaction.
provides the energy and drives the fusion of
viral and endosomal membranes. Further An important question in the context of TBEV
85 fusion relates to the molecular switches that
biochemical studies indicated that the
structural conversion of E was a two-step sense the acidic pH in endosomes and induce
process, in which the acidic pH in endosomes the fusogenic conformational change in E.
first caused the dissociation of E dimers Because of their pKa near the pH threshold of
followed by an irreversible trimerization. fusion, hisitidines have been hypothesized to
Monoclonal antibody studies and mutational play such a role in the fusion trigger. There are
analyses provided evidence that the highly indeed five histidines (H146, 248, 287, 323, 438)
conserved sequence element in E, located at that are absolutely conserved among flavivirus E
the tip of DII and now designated fusion loop proteins, suggesting an indispensable structural
(FL), was responsible for interacting with the and/or functional role in the viral life cycles. The
endosomal target membrane as an initial step in use of RSPs (see above) with mutated histidines
membrane fusion. 55,79 at these positions allowed the identification of
H323 as a key residue for triggering the acidic-
It was a key finding of these studies that the pH-induced trimerization of E and concomitant
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soluble E dimer (which dissociates into membrane fusion. This residue is involved in
monomers at acidic pH) could be converted into intramolecular interactions at the interface
a trimer in the presence of liposomes, 79,86 laying between DI and DIII in the E dimer. Its protona-
the foundation for the crystallization of this post tion apparently facilitates E dimer dissociation
-fusion conformation and the determination of and allows DIII to be released from its original
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its atomic structure by X-ray crystallography position and to relocate as required for post-
(Figure 3). The structure revealed that the fusion trimer formation (Figure 3). Other
folding of the three domains is maintained but conserved histidines were shown to be
that their relative orientation is altered (Figure dispensable for fusion, but they may have
3A,B). Specifically, DIII relocates from its critical roles in unrelated low-pH-driven
position at the end of the dimer to the side of processes of the viral life cycle, such as virus
the trimer in such a way that a hairpin-like maturation (Figure 2).
structure is formed (Figure 3A), in which the FL

and the stem-anchor region of E would be
juxtaposed in the full-length E trimer. This Antigenic structure of TBEV and
structure was reminiscent of the post-fusion
structures of class 1 viral fusion proteins such as virus neutralization
the influenza virus hemagglutinin and suggested
– in combination with studies on fusion Because of its functions in flavivirus attachment
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intermediates – that the TBEV fusion and entry as well as membrane fusion in
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