# Structural basis for HIV-1 Gag interactions with cellular and viral constituents

> **NIH NIH R01** · UNIVERSITY OF ALABAMA AT BIRMINGHAM · 2022 · $456,522

## Abstract

During the late phase of HIV-1 infection, the Gag polyproteins are transported to the plasma membrane (PM) for
assembly. Gag targeting and assembly on the PM is dependent on specific interactions between its matrix (MA)
domain and phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), a signaling lipid localized on the inner leaflet of
the PM. Concurrent or subsequent to Gag assembly, the envelope (Env) protein is recruited to the PM for
incorporation into virus particles, a process that is dependent on Gag assembly. Several lines of evidence
suggest that incorporation of Env is mediated by interactions between the MA domain of Gag and the cytoplasmic
tail of gp41 (gp41CT), a mechanism that remains to be elucidated. For over a decade, we have pioneered
approaches to investigate at the molecular level how retroviral (HIV-1, HIV-2, MLV, and ASV) MA domains of
Gag interact with lipids (e.g., PI(4,5)P2) and membranes, a key requirement for understanding Gag assembly
and ultimately Env incorporation. A major barrier to characterizing gp41CT–MA interaction has been the
unavailability of a recombinant gp41CT protein and the inability to reconstitute the proper conditions for the
interaction to occur. During this funding period we have recorded a breakthrough by determining the three-
dimensional structure of HIV-1 gp41CT and characterized its interaction with membrane. This achievement, an
elusive task for over two decades, filled a major gap by providing the structure of the last segment of HIV-1
proteome. In this renewal, we set our sights on elucidating the molecular mechanism by which Gag mediates
the recruitment of HIV-1 Env into assembly sites. We will test the hypothesis that trimerization of the MA domain
plays an important role in gp41CT interaction and therefore Env recruitment into particles. We will employ a
battery of structural biology, biophysical, and biochemical tools to generate a macromolecular picture of how the
MA domain of Gag binds to membrane and how it interacts with gp41CT. Our specific aims are: (i) Engineer
HIV-1 MA trimer and hexamer and characterize their binding to membranes, (ii) characterize the interactions
between HIV-1 MA and gp41CT, and (iii) determine the structure of MA–gp41(TM-CT)–membrane complex by
single-particle cryo-EM. This proposal rests on a solid premise of: (1) 25 years of functional data that is waiting
for a molecular interpretation to provide sharper insights into Gag assembly on the PM and mechanisms of Env
incorporation, (2) highly technical skill sets for working with proteins, membranes, structural and biophysical
tools, (3) strong preliminary data, and (4) an outstanding collaborative team with 20-30 years of experience in
cryo-EM, x-ray crystallography, and mass spectrometry. Therefore, the proposed structural studies will provide
a detailed understanding of HIV-1 Gag assembly on the PM and Gag–mediated Env incorporation. We hope that
the outcome of this proposal will help in the development of new ...

## Key facts

- **NIH application ID:** 10462579
- **Project number:** 5R01AI150901-13
- **Recipient organization:** UNIVERSITY OF ALABAMA AT BIRMINGHAM
- **Principal Investigator:** Jamil Subhi Saad
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $456,522
- **Award type:** 5
- **Project period:** 2010-05-15 → 2023-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10462579

## Citation

> US National Institutes of Health, RePORTER application 10462579, Structural basis for HIV-1 Gag interactions with cellular and viral constituents (5R01AI150901-13). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10462579. Licensed CC0.

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