A novel method for viral protein tracking in host cells

Barry, Jacqueline Patricia (2017) A novel method for viral protein tracking in host cells. Masters thesis, Memorial University of Newfoundland.

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Abstract

Hepatitis C Virus (HCV) infects approximately 70 million people worldwide and chronic infection can lead to liver cirrhosis and hepatocellular carcinoma. HCV contains a perplexing protein that has a number of proposed functions. This protein, termed p7, is essential for virus infectivity in vivo, however its function is a matter of controversy. Research into the function of p7 has been limited because there are no reliable antibodies available for the visualization of this protein. The goal of this project was to establish a system that utilizes fluorescent unnatural amino acids in order to label p7 within the context of a replicating virus. Strategically placed mutations within the viral p7 protein were selected to test their amenability to incorporation of an unnatural amino acid. In order to optimize the incorporation system, plasmids containing mutations in the viral core protein were synthesized for further screening of positions tolerable to substitution. Ultimately, we were successful in incorporating a fluorescent unnatural amino acid into the viral core protein in a single protein expression system. If we can successfully transfer this system back into the context of a replicating virus, this technique could be used to facilitate the study of viral proteins in HCV and other viruses.

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