An Absolutely Conserved Tryptophan in the Stem of the Envelope Protein E of Flaviviruses Is Essential for the Formation of Stable Particles

Abstract

The major envelope protein E of flaviviruses contains an ectodomain that is connected to the transmembrane domain by the so-called “stem” region. In mature flavivirus particles, the stem is composed of two or three mostly amphipathic α-helices and a conserved sequence element (CS) with an undefined role in the viral life cycle. A tryptophan is the only residue within this region which is not only conserved in all vector-borne flaviviruses, but also in the group with no known vector. We investigated the importance of this residue in different stages of the viral life cycle by a mutagenesis-based approach using tick-borne encephalitis virus (TBEV). Replacing W421 by alanine or histidine strongly reduced the release of infectious virions and their thermostability, whereas fusion-related entry functions and virus maturation were still intact. Serial passaging of the mutants led to the emergence of a same-site compensatory mutation to leucine that largely restored these properties of the wildtype. The conserved tryptophan in CS (or another big hydrophobic amino acid at the same position) is thus essential for the assembly and infectivity of flaviviruses by being part of a network required for conferring stability to infectious particles.

Molecular Organisation of Tick-Borne Encephalitis Virus

“…Specific deformation of the membrane curvature at these points (Figure 1) indicates that lipid rafts are probably important in assembly [58,59]. The lipid pocket-forming residues Arg412, Trp421, His438, and Met490 of E play critical role in particle formation as shown in mutagenesis studies in ZIKV, DENV, SPONV, and TBEV [16,17,19,60]. However, the functions of Thr406, Ile410 in E and Thr13 in M have not been elucidated.…”