# Single-particle analysis of HSV-1 membrane fusion mechanism

> **NIH NIH R21** · TUFTS UNIVERSITY BOSTON · 2020 · $253,583

## Abstract

PROJECT SUMMARY/ABSTRACT
Herpes Simplex virus type 1 (HSV-1) requires four glycoproteins for cell entry and membrane fusion – gB, gD,
gH, and gL – in addition to a cellular receptor. This number far exceeds the number of glycoproteins utilized by
most other enveloped viruses complicating mechanistic studies of HSV-1-mediated entry and fusion. We now
know that in HSV-1, as in other herpesviruses, the receptor-binding, regulatory, and fusogenic functions are
distributed among several glycoproteins. gB is the conserved fusogen that, by analogy with other viral
fusogens, is thought to facilitate membrane merger by undergoing large-scale refolding that brings the viral and
cellular membranes together. However, gB is unusual in requiring several additional proteins for function.
Therefore, understanding how gB works to mediate fusion during cell entry requires direct measurements of its
fusogenic activity. Recently, single-particle tracking (SPT) has emerged as a powerful tool for quantitative
studies of fusion of individual virions with fluid, supported lipid bilayers using total internal reflection microscopy.
The SPT approach enables direct visualization and kinetic measurements of the viral fusion pathway and has
been successfully used with both pH-triggered and receptor-triggered fusogens. The goal of this exploratory
proposal is to develop single-particle imaging of HSV-1 fusion with supported lipid bilayers to visualize different
stages in fusion, measure their kinetic parameters, identify kinetic intermediates, and correlate them with
structural rearrangements in gB. Given that the SPT approach has not yet been applied to viruses that utilize
>1 glycoprotein and may be challenging to apply to HSV-1 that has 15 envelope proteins, Aim 1 will utilize
Vesicular Stomatitis Virus (VSV) virions lacking the native fusogen G and pseudotyped with HSV-1 entry
glycoproteins gB, gD, gH, and gL (VSVDG-BHLD), which retain key characteristics of HSV-1 entry. In Aim 2,
the SPT approach will be extended to HSV-1, a more complicated yet more biologically relevant system. The
scientific premise of the proposed work is that harnessing the power of single-particle imaging of virion fusion
provides a unique opportunity to address the lingering questions in HSV-mediated membrane fusion
mechanism, with the ultimate goal of reconstructing the HSV-1-mediated fusion pathway more fully.

## Key facts

- **NIH application ID:** 9995909
- **Project number:** 1R21AI145272-01A1
- **Recipient organization:** TUFTS UNIVERSITY BOSTON
- **Principal Investigator:** Ekaterina Heldwein
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $253,583
- **Award type:** 1
- **Project period:** 2020-09-03 → 2022-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9995909

## Citation

> US National Institutes of Health, RePORTER application 9995909, Single-particle analysis of HSV-1 membrane fusion mechanism (1R21AI145272-01A1). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9995909. Licensed CC0.

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