# Structural and mechanistic basis of AAGAB-controlled AP2 adaptor assembly

> **NIH NIH R01** · FLORIDA STATE UNIVERSITY · 2021 · $344,202

## Abstract

PROJECT SUMMARY
In clathrin-mediated endocytosis (CME), the coat protein clathrin relies on adaptors to recruit cargo proteins to
endocytic sites. The predominant clathrin adaptor in CME is the heterotetrameric AP2 adaptor complex, which
is comprised of two large subunits ( and β), one medium subunit (µ), and one small subunit (). It is generally
thought that multimeric trafficking adaptors such as AP2 adaptor assemble spontaneously. However, we recently
discovered that AP2 adaptor assembly is an ordered process controlled by alpha and gamma adaptin binding
protein (AAGAB). Without the assistance of AAGAB, AP2 adaptor fails to form, leading to CME defects. These
findings revealed a previously unrecognized pathway in clathrin-mediated trafficking. However, it remains
unclear how AAGAB recognizes AP2 subunits and how AAGAB-AP2 interactions drive AP2 adaptor assembly.
In our preliminary studies, we expressed and purified free AAGAB and AAGAB:AP2 assembly intermediates,
and gained initial insights into their structures and functions. In this research, we will take advantage of these
preliminary data to determine the structural and mechanistic basis of AAGAB-controlled AP2 adaptor assembly.
We will first characterize the biochemical properties of free AAGAB and AAGAB:AP2 assembly intermediates
using reconstituted systems. We will then determine the atomic structures of the proteins and protein complexes
using X-ray crystallography and single-particle cryo-electron microscopy. Next, we will validate the physiological
relevance of the biochemical and structural findings using cell-based genetic assays. Finally, we will determine
how AAGAB functions are impacted by disease-causing mutations. Successful completion of this proposed
research will fill a major gap in our knowledge of membrane protein trafficking. This work will also serve as a
paradigm for understanding the assembly of trafficking adaptors in general. Ultimately, these findings will
facilitate the development of novel therapeutic strategies for human diseases caused by AAGAB mutations.

## Key facts

- **NIH application ID:** 10215568
- **Project number:** 5R01GM138685-02
- **Recipient organization:** FLORIDA STATE UNIVERSITY
- **Principal Investigator:** Qian Yin
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $344,202
- **Award type:** 5
- **Project period:** 2020-07-13 → 2024-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10215568

## Citation

> US National Institutes of Health, RePORTER application 10215568, Structural and mechanistic basis of AAGAB-controlled AP2 adaptor assembly (5R01GM138685-02). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10215568. Licensed CC0.

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