All ETDs from UAB

Advisory Committee Chair

Jamil S Saad

Advisory Committee Members

Terje Dokland

Todd J Green

Chad M Petit

Peter E Prevelige

Document Type

Dissertation

Date of Award

2020

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Recent advancements in drug therapies and patient care have drastically improved upon the mortality rates of HIV-1 infected individuals. Many of these therapies were developed or improved upon using structure-based techniques, which underscores the importance of obtaining atomic level structural data critical to the understanding of essential mechanisms in the replication cycle of HIV-1. One such process which remains poorly understood on the structural level is the incorporation of the envelope glycoprotein (Env) into budding virus particles. The Env protein mediates viral infection of host cells, and there is strong evidence suggesting that incorporation of Env is mediated by an interaction, localized on the plasma membrane (PM), between its own cytoplasmic domain (gp41CT) and the Matrix domain (MA) of the Gag polyprotein. However, there are currently no structural or biophysical data which describe this interaction in detail, largely due to difficulties in reconstituting the proteins in a membrane-like environment suitable for structural investigations. In Chapters 2 and 3 of this dissertation, I discuss my efforts to independently characterize the structures and membrane interactions of gp41CT and MA respectively. Gp41CT represents the last unsolved structure in the HIV-1 proteome. In Chapter 2, I determined the solution structure and characterized the membrane interactions of gp41CT using NMR techniques. These studies allowed me to resolve longstanding uncertainties about the overall topology of gp41CT and provided insights into how the gp41CT protein facilitates interactions with its multiple cellular binding partners and mediates Env incorporation. In Chapter 3, I employed various biophysical and structural techniques to better understand the structure and membrane interactions of the trimeric MA protein. MA has been shown to exist in a monomer/trimer equilibrium in solution, and evidence suggests that the trimerization of MA on the membrane is directly linked to Env incorporation. Herein, I provide new evidence for the location of the MA-MA interface involved in trimerization and characterize the role of trimerization in membrane binding. The methods and results presented here advance our understanding of key mechanisms in HIV-1 assembly and provide a template for further structural investigations into the interaction between MA and gp41CT.

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