# Structural and Mechanistic Insights into AAV Rep Mediated Site-Specific Integration and Packaging

> **NIH NIH R01** · VIRGINIA COMMONWEALTH UNIVERSITY · 2022 · $92,451

## Abstract

PROJECT SUMMARY
Adeno-Associated Virus (AAV) has evolved the property to integrate into its host genome exploiting a unique
mechanism shared by organisms in all domains of life to process single stranded DNA during DNA replication,
bacterial conjugation and special cases of transposition. The large AAV Rep proteins Rep68/Rep78 are the
key players that carry out this process using two multifunctional domains that include an origin binding domain
(OBD) that binds DNA specifically and belongs to the family of HUH endonucleases; and a SF3 helicase
domain. Integration is contingent on the binding of Rep68/Rep78 to the integration site AAVS1, promote its
melting and perform a strand-specific transesterification reaction in the single-stranded region that has been
extruded during the melting step. Little is known about the molecular details of this process and our long-term
goal is to determine the molecular mechanism of site-specific integration mediated by AAV Rep proteins. In
this proposal we will focus on uncovering the mechanisms of Rep mediated DNA melting, nicking and
packaging using a combination of structural biology methods that include X-ray crystallography, and single-
particle reconstruction Cryo-EM coupled to biochemical, biophysical and in vivo assays.

## Key facts

- **NIH application ID:** 10581772
- **Project number:** 3R01GM124204-05A1S1
- **Recipient organization:** VIRGINIA COMMONWEALTH UNIVERSITY
- **Principal Investigator:** Carlos R Escalante
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $92,451
- **Award type:** 3
- **Project period:** 2017-09-30 → 2025-11-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10581772

## Citation

> US National Institutes of Health, RePORTER application 10581772, Structural and Mechanistic Insights into AAV Rep Mediated Site-Specific Integration and Packaging (3R01GM124204-05A1S1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10581772. Licensed CC0.

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