Single-Cell Biophysics: Measurement, Modulation, and Modeling

Single-Cell Biophysics: Measurement, Modulation, and Modeling

Poster Abstracts

34-POS Board 17 Integrative Models of Complete Viruses – Combining Structure, Genomics and Modeling Roland G. Huber 1 , Jan K. Marzinek 1,2 , Daniel Holdbrook 1 , Ana de Suoto Martins 3 , Ivo Martins 3 , Yue Wan 4 , Peter J. Bond 1,2 . 1 A*STAR Bioinformatics Institute (BII), Singapore, Singapore, 2 NUS Department of Biological Sciences (DBS), Singapore, Singapore, 4 A*STAR Genome Institute Singapore (GIS), Singapore, Singapore. 3 Instituto de Medicina Molecular, Lisboa, Portugal, Dengue and Zika viruses belong to the Flavivirus family of mosquito-borne infections prevalent in tropical and subtropical regions. They are enveloped, single-stranded positive-sense RNA viruses with a genome of approximately 11kb and form virions of ~45 nm in diameter. In recent years, near-atomistic cryo-EM structures have become available. While these structures reveal the precise organization of the protein envelope, they are not able to resolve either the lipid membrane or the viral core in detail. To probe deeper into the viral particle, we created detailed models of the viral envelope including the lipid vesicle circumscribed by the envelope proteins. These models reveal a tensioned membrane structure that follows the icosahedral pattern given by the symmetry of the surrounding proteins. We show how anionic lipids within the lipid vesicle anchor the membrane to the protein coat. We propose that this tension within the deformed lipid vesicle may store energy that is used to overcome the activation barrier for fusion with the late endosomal membrane and subsequent release of the viral genome. Within the envelope, the viral genome interacts with a highly charged capsid protein. Cryo-EM structures do not show a simple geometric capsid as commonly found in non-enveloped viruses. In order to determine interaction sites of this capsid protein with the viral genome we performed a nuclease digestion assay: protein-bound segments are better protected from digestion and can be sequenced. Subsequent alignment against the full genome reveals the location of these protected fragments. Integrating structural, biochemical, genomic, and computational methods thus allowed us to create a near-atomistic, dynamic model of the mature virion. We are currently modeling the fusion of these particles with endosomal membrane models, which may allow us to identify novel targets for therapeutics against these and other Flaviviruses.

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