Dynamics and evolution of HIV GAG and gagpol processing

This Project Grant award, provided by the National Institute of Allergy and Infectious Diseases (NIAID) under the Allergy and Infectious Diseases Research program (CFDA 93.855), supports research to investigate the dynamics and evolution of HIV Gag and GagPol processing. The $752,745 award aims to use molecular dynamics simulations, single-molecule FRET imaging, and cryo-electron tomography to probe the structure and dynamics of Gag and GagPol that enable ordered processing by the viral protease. This research will help elucidate the evolutionary constraints on Gag and GagPol that ensure sufficient dynamics for efficient virus maturation, and inform the development of the next generation of HIV maturation inhibitors. The award period runs from August 6, 2024 to July 31, 2025 and is being administered by the University of Massachusetts Medical School, a prominent research institution with extensive experience in federal grants and contracts.

Abstract

Maturation of the HIV-1 capsid requires activation of the viral protease (PR) and proteolytic processing of the GAG and gagpol polyproteins. Previous studies support a model in which HIV-1 maturation depends on the proper order and timing of proteolytic events in GAG by PR. According to this model, ordered GAG processing is determined by the rate of proteolysis at the individual cleavage sites. The structural and dynamic features of GAG, gagpol, and the immature virion that determine the kinetics and order of processing by PR, including the autoprocessing of gagpol, have not been characterized. Furthermore, the evolutionary constraints that ensure the maintenance of ordered and efficient GAG processing are unknown. Previous studies of GAG processing and gagpol autoprocessing have been limited by a lack of experimental approaches capable of probing dynamic events and the organization of gagpol within virions. Furthermore, molecular simulations have generally been performed on reduced structures that do not capture the full complexity of GAG and gagpol complexes. Here, we will use molecular dynamics (MD) simulation of full-length GAG and gagpol complexes to probe the dynamics that occur prior to and during interaction with PR and provide atomic resolution detail of these interactions. These simulations will suggest hypotheses regarding large-scale motions of GAG and gagpol domains that will be tested through the application of single-molecule förster resonance energy transfer (smfret) imaging and cryo- electron tomography (cryoet). These approaches enable visualization of the structure and dynamics of GAG and gagpol that enable ordered processing by PR in the context of polyprotein monomers, dimers, hexamers, and intact immature virions. Finally, through consideration of diverse HIV-1 strains and GAG variants that circumvent inhibition of maturation by modulating the flexibility of the immature lattice, we will determine the evolutionary constraints on GAG and gagpol that ensure sufficient dynamics are maintained. These data will inform development of the next generation of maturation inhibitors.