Viruses are biological entities that constantly affect the world around us and in many cases they can harm the health of humans and negatively impact food production, in agriculture andfisheries. The project concept is that, through the structural studies of viruses that infectsimple hosts, it is possible to point out structural features that are unique and conserved inviruses, that infect human, other animals, and crops, in the same phylogenetic lineage. Theseunique features have functionally been acquired, and are therefore potential drug and vaccinetargets. Standing on this concept, we aim to study three groups of pathogenic viruses: (i)Protozoan/Yeast Totiviridae and metazoan totivirus-like viruses, (ii) Invertebrate flavi-likeand vertebrate Flaviviridae viruses, and (iii) Invertebrate Mesoniviridae and vertebrateCoronaviridae viruses.First, we focused on acquired capsid structures of the totivirus-like Omono River Virus(OmRV). Unlike yeast Totiviridae viruses, the totivirus-like OmRV capsid presents surfaceprotrusion proteins, which are expected to be crucial for its extracellular transmission. Theinteraction between the capsid and the protrusion proteins is hypothesized to be largelymediated by amino acid residue T365 of the protrusion proteins. To clarify the function ofthese protrusion proteins we have successfully developed molecular tools, which are arecombinantly expressed protrusion protein and an OmRV infectious DNA clone with aT365A mutation. Although the excess of the protrusion proteins does not remarkably affectthe infectivity and binding capabilities of the OmRV particles, the OmRV with the T365Amutation shows a significantly reduced infectivity. Assuming that the infectivity was partiallyeliminated by the mutation, a putative transmission mechanism is associated with theprotrusion proteins. However, it is still under debate whether the protrusion proteins are anessential factor for extracellular transmission. Additionally, a new method was preliminarilyused for observing the interaction between the OmRV capsid and protrusion proteins usingdifferential scanning fluorimetry. The findings and the established methods can contribute tothe understanding of molecular mechanisms present in other pathogenic totivirus-like virusesthat affect fish, such as the salmon piscine myocarditis virus (PMCV).Secondly, we focused on acquired surface structures of vertebrate Flaviviridae andCoronaviridae viruses. Unlike the invertebrate flavi-like and Mesoniviridae viruses, thevertebrate viruses have acquired surface structures that are expected to be critical for evadinghosts’ adaptive immunity. To discover these acquired surface structure, it is necessary todetermine the surface structure of the invertebrate flavi-like and Mesoniviridae viruses. Herewe report the successful expression and purification of the invertebrate flavi-like Southernpygmy squid flavivirus (SpsFV) precursor/membrane-envelope (prM-E) protein and theMesoniviridae Nam Dinh virus (NDiV) p2a spike protein. The established expression andpurification system can be further used to resolve their structures using cryo-EM singleparticle analysis in future studies.
| Identifer | oai:union.ndltd.org:UPSALLA1/oai:DiVA.org:uu-470848 |
| Date | January 2022 |
| Creators | Filipe, Diogo |
| Publisher | Uppsala universitet, Institutionen för biologisk grundutbildning |
| Source Sets | DiVA Archive at Upsalla University |
| Language | English |
| Detected Language | English |
| Type | Student thesis, info:eu-repo/semantics/bachelorThesis, text |
| Format | application/pdf |
| Rights | info:eu-repo/semantics/openAccess |
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