# Massively Parallel Mutational Analyses of Viral Neutralization Mechanisms Against Human Restriction Factors

> **NIH NIH K22** · CASE WESTERN RESERVE UNIVERSITY · 2020 · $108,000

## Abstract

PROJECT SUMMARY / ABSTRACT:
 Viral pathogens remain a major public health threat, and their persistence is largely due to the
mechanisms they have adapted to evade or combat our immune systems. Cellular defense proteins called
restriction factors are important for stifling the initial spread of infection both within and between people, but
many viruses have evolved proteins capable of antagonizing them. The protein surfaces used by many viral
proteins to bind and inactivate restriction factors are not known, making it difficult to disrupt them to promote
immunity. Mutagenesis experiments can address these gaps in our knowledge, but conventional methods are
too slow and laborious to test the many thousands of possibly informative protein mutants that exist.
 I propose to use a new experimental approach called deep mutational scanning to study how all of the
possible single amino acid variants of a viral immune evasion protein or host restriction factor affects the ability
of a virus to infect cells. In the first aim, I will test thousands of protein variants of the HIV-1 protein Vif to figure
out which amino acids it uses to bind and neutralize each of the different APOBEC3 restriction factors. This will
help us understand how Vif has evolved to keep binding each APOBEC3 protein without losing binding against
the others. This work will also help us understand and improve upon antivirals that target Vif. In the second
aim, I will assay the activity of thousands of variants of the restriction factor Tetherin in the presence of diverse
viral proteins that have evolved to neutralize it. By comprehensively identifying Tetherin variants capable of
escaping each antagonist, I will infer each of their binding sites. By comparing the relative strength and number
of possible escape variants for each antagonist, I will learn which mechanisms are most effective in
neutralizing Tetherin.
 This work will reveal discrete proteins interaction surfaces that may be disrupted by antiviral drugs.
These results will also reveal successful strategies viruses have evolved to inhibit immune proteins, which will
make us better able to find new ways to stop them. This project will highlight the power of using deep
mutational scanning to gain broad, unbiased views into the mechanisms that viruses have evolved to
neutralize our immune systems, which I plan to expand upon in my research lab.

## Key facts

- **NIH application ID:** 9990669
- **Project number:** 5K22AI141620-02
- **Recipient organization:** CASE WESTERN RESERVE UNIVERSITY
- **Principal Investigator:** Kenneth A Matreyek
- **Activity code:** K22 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $108,000
- **Award type:** 5
- **Project period:** 2019-08-07 → 2021-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9990669

## Citation

> US National Institutes of Health, RePORTER application 9990669, Massively Parallel Mutational Analyses of Viral Neutralization Mechanisms Against Human Restriction Factors (5K22AI141620-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9990669. Licensed CC0.

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