Optimized Microbial Recombinant Production of HIV-1 Anti-Envelope Antibody Fragments with Applications to Single Particle Tracking of Virus Assembly

<p>In my findings, I have established a set series of protocols to recombinantly produce, purify and apply various fluorescent probes in vitro for the fluorescent labeling and study of the human immunodeficiency virus type 1 (HIV-1) envelope (Env) protein during HIV viral assembly. There remains insufficient knowledge about the molecular dynamics and interactions of HIV-1 Env protein with its counterpart, Gag, on the inner host cell surface during assembly of a mature virus particle. There also remains an insufficient amount of data for the understanding and clarification of the mechanism of action of a known host cell HIV-1 restriction factor, Serinc5 (SER 5), which is believed to have correlation with Env in fusion events.</p> <p>Using superresolution microscopy to measure and track host–pathogen interfaces that occur on a scale below the resolution limit of the light microscope is becoming increasingly popular. I have used total internal reflection microscopy (TIRF) and Photoactivated Localization Microscopy (PALM) to depict sites of molecular assembly via Env and Gag colocalization as well as measure and track other means of Env such as diffusion rates and host cell restriction factor interactions.</p> <p>Overall, my findings have provided improved strategies and tools for the use in fluorescently labeling and tracking HIV-1 Env and an HIV-1 host cell restriction factor known as SER 5. Superresolution microscopy and single particle tracking can be achieved provided high specificity, brightness and purity while maintaining low KDs. The production and data of these probes are imperative for both understanding the pathological processes of HIV-1 assembly and other stages of the HIV-1 life cycle as well as the various protein-protein interactions.</p>

Name thesis:degree_name

M.S.

Level thesis:degree_level

Masters Thesis

Year dc:date.available

2021

Author dc:creator

• Bruns, Merissa Michelle

Contributors dc:contributor

• Schuyler Van Engelenburg
• Michelle Knowles
• Erich Kushner
• Cedric Asensio

• HIV-1
• Superresolution microscopy
• Biochemistry, Biophysics, and Structural Biology
• Biology
• Biophysics
• Immunology and Infectious Disease
• Life Sciences
• Microbiology
• Virology

Statement dc:rights

• <p>Copyright is held by the author. User is responsible for all copyright compliance.</p>

Language dc:language

en

Repository record dc:identifier

https://digitalcommons.du.edu/etd/1872

OAI identifier oai:identifier

oai:digitalcommons.du.edu:etd-2865