Event Title

Oligomerization of Hepatitis C Virus Helicase Using Recombinant Fusion Proteins

Mentor 1

Dr. David Frick

Location

Union Wisconsin Room

Start Date

29-4-2016 1:30 PM

End Date

29-4-2016 3:30 PM

Description

Hepatitis C is a blood-borne RNA virus, which employs host cell machinery to undergo viral replication. A core component needed for HCV replication is NS3, or Non-Structural Protein 3. This protein consists of an N-terminal and a C-terminal domain. The N-terminal domain containing a protease activity, and the C-terminal domain a helicase activity. Our objective is to study the helicase activity of NS3, in order to better understand whether it functions as a monomer, dimer, or higher order oligomer. We hope to achieve this through the use of recombinant fusion proteins, with which we will fluorescently label domains of interest on NS3. The fluorophores used being CFP and YFP tagged NS3. This fluorescence can then be analyzed using F.R.E.T (Fluorescent Resonance Energy Transfer). F.R.E.T. allows for monitoring of energy transfer between a donor and acceptor fluorophore, and the degree of transfer can be analyzed to determine the proximity between the two fluorescently tagged NS3h proteins. The separately tagged proteins will be analyzed with F.R.E.T. under varying DNA concentrations, to gain insight into whether or not multiple NS3 proteins come together to express helicase activity.

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Apr 29th, 1:30 PM Apr 29th, 3:30 PM

Oligomerization of Hepatitis C Virus Helicase Using Recombinant Fusion Proteins

Union Wisconsin Room

Hepatitis C is a blood-borne RNA virus, which employs host cell machinery to undergo viral replication. A core component needed for HCV replication is NS3, or Non-Structural Protein 3. This protein consists of an N-terminal and a C-terminal domain. The N-terminal domain containing a protease activity, and the C-terminal domain a helicase activity. Our objective is to study the helicase activity of NS3, in order to better understand whether it functions as a monomer, dimer, or higher order oligomer. We hope to achieve this through the use of recombinant fusion proteins, with which we will fluorescently label domains of interest on NS3. The fluorophores used being CFP and YFP tagged NS3. This fluorescence can then be analyzed using F.R.E.T (Fluorescent Resonance Energy Transfer). F.R.E.T. allows for monitoring of energy transfer between a donor and acceptor fluorophore, and the degree of transfer can be analyzed to determine the proximity between the two fluorescently tagged NS3h proteins. The separately tagged proteins will be analyzed with F.R.E.T. under varying DNA concentrations, to gain insight into whether or not multiple NS3 proteins come together to express helicase activity.