Date of Award

12-10-2018

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Chemistry

First Advisor

Dr. Ming Luo

Second Advisor

Dr. Jenny Yang

Third Advisor

Dr. Gregory Poon

Abstract

Understanding the basic structure-function relationships that guide the biology of various pathogenic viruses is the cornerstone in the development of antiviral therapies. One group of highly pathogenic viruses, which desperately needs novel antiviral therapies, are Negative Strand RNA viruses. This group of viruses contains some of the most highly pathogenic and pandemic viruses known: like Ebola, Influenza, Marburg, and Rabies viruses. Between 2013 and 2016, during the latest endemic outbreak of Ebola, the World Health Organization estimated that it caused over 11,000 deaths. However, without in-depth understanding of the basic science that guides the replication of these viruses, design of targeted therapies is hard. Utilizing the model virus for Negative Strand RNA viruses, Vesicular Stomatitis virus, this work studies the structure-function relationships of the nucleocapsid protein. Since the nucleocapsid encapsulates the virus’ genomic material this protein is increasingly important study. Furthermore, the virus cannot undergo replication if the polymerase does not recognize the nucleocapsid for viral RNA synthesis. This work takes a three-pronged approach when investigating the nucleocapsid: development of a novel antiviral to target the nucleocapsid, the use of structural information to study the possible interactions between the nucleocapsid and the polymerase, and a study of the genomic constraints placed on polymerase activity by the nucleocapsid. Utilizing an already characterized class of compounds, polyamides, we showed that it is possible to target the genomic RNA in the nucleocapsid of a Negative Strand RNA virus and inhibit viral replication. This is the first known instance of this occurring in Negative Strand RNA viruses. To examine the structural interactions between the nucleocapsid and the polymerase a flexible loop was hypothesized to be integral toward the formation of an active polymerase complex. Alanine scanning deletions yielded an abolishment in polymerase activity, which could be restored after compensatory mutations were allowed to occur. Finally, the nucleotide content sequestered within the nucleocapsid has a direct effect on the processivity of the polymerase complex, and ushers in a paradigm changing model for how one examines Negative Strand RNA virus activity and the evolutionary constraints placed on the genomic RNA. Merging the findings of these studies together, understanding the structure-function relationships of the nucleocapsid can yield a lot of knowledge about the viral life-cycle of the entire Negative Stranded RNA virus family.

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