Structures of flavivirus particles

My Ph.D research involves the study of flavivirus particle structures. Three structural proteins, E, prM/M and C, exist in the flavivirus particles. The Capsid (C) protein is important in the genome packing. Both the premembrane (prM) and envelope (E) proteins are transmembrane glycoproteins. PrM is cleaved during maturation, leaving only M in the now infectious particles. E participates in binding to cellular receptors and interacting with host cell membranes for fusion. Structures of immature flaviviruses have been determined to 12.5Å resolution with cryo EM image reconstruction techniques. The structure showed 60 prominent icosahedrally packed trimeric spikes on the viral surface. Each spike is composed of three asymmetric prM-E heterodimers. The prM protein covers the fusion peptide of the associated E protein, preventing E from losing the fusogenic activity prior to maturation. E and prM are arranged in a similar manner as that of the E1 and E2 proteins in alphaviruses. The crystal structure of the E ectodomain (sE) was determined by X-ray crystallography. By comparing it with a previously determined crystal structure, it was found that there is a 10° rotation about a hinge relating domains DI and DII. The crystal structure of sE was then divided into two rigid bodies (DI + III, DII), which were then independently fitted into EM maps of both mature and immature virus particles. It was shown that there was a rotation of 27° between the two rigid bodies about the same hinge observed when comparing different crystal structures. The flexibility is apparently a functional requirement for assembly of flaviviruses. The structure of the nucleocapsid core was poorly observed in the reconstructions of flavivirus particles. This could be caused by the domination of the glycoproteins in the orientation search of each particle in the reconstruction. Thus, in order to determine the structure of the core, the density contributed from the glycoprotein layer was subtracted from the 2D raw images of whole virus particles. The reconstruction of the core was then performed from the difference images. It was demonstrated that the nucleocapsid core of flaviviruses does not have a unique structure, in agreement with some biological observations