EPSRC-SFI: Understanding Molecular Interactions Initiating Adsorption of Viral Capsid Proteins on Lipid Droplets (MIrACLe)

Dengue virus (DENV), a member of the flaviviridae family, is the cause of mosquito-borne dengue fever. Endemic in 100 countries, it leads to over 400 million infections and 25k deaths annually worldwide. With no effective treatments or vaccines, it represents a major socio-economic burden for tropical and subtropical developing countries. Expansion of dengue Aedes mosquito vector due to climate change makes it increasingly a global threat, with over 4 billion people at risk, including EU and UK. There is an urgent need for detailed understanding of the DENV lifecycle for devising new pathways for tackling DENV infections. Central to DENV's lifecycle is its replication, a process that has been very recently linked with lipid droplets (LD; ubiquitous multifunctional intracellular organelles of 0.05-100 um in size), with DENV capsid proteins (DENV-C) found accumulating on LD, leading to nucleocapsid formation and viral particle self-assembly. Critically, this process is initiated by the adsorption of DENV-C onto the LD surface mediated by molecular interactions between DENV-C and the LD membrane and surface-anchored proteins, particularly Perilipin 3 (Plin3). Of particular interest to the molecular interactions is an intrinsically disordered region (IDR), i.e. the first 30 amino acid residues at the DENV-C N-terminus. Probing these fundamental molecular interactions, using a combination of physicochemical experimental and computer simulation methods, is the focus of this proposal. Our approach to designing the experimental programme is guided by the following considerations. The molecular interactions - electrostatic or hydrophobic - depend intricately on the amino acid sequence in DENV-C IDR. We thus will leverage advanced peptide synthesis to precisely tailor IDR-analogous viral peptide sequences and compare them with full IDR and C-proteins. The inherent complexity in LD obscures mechanistic probing of the underpinning molecular interactions. We thus will leverage biophysical methods to establish LD models, incorporating Plin3 and essential LD surface structural and compositional features. Going beyond phenomenological observations, we will bring to bear quantitative experimental and computational methods in biophysics and surface science to directly access molecular structures and interactions at complex interfaces.