# Folding and Chaperone Interactions of Multi-domain Proteins

> **NIH NIH R01** · JOHNS HOPKINS UNIVERSITY · 2024 · $149,773

## Abstract

Project Summary
Combining several functional units, termed domains, into a single polypeptide chain is a common
evolutionary strategy for creating biological complexity. The resulting multi-domain proteins are
prevalent in all proteomes and carry out essential cellular functions. However, the increased functional
complexity of these large proteins complicates their folding into native functional structures. In contrast
to many smaller proteins or individual domains, multi-domain proteins are prone to misfolding and
potentially cytotoxic aggregation. In the cell, several factors ensure efficient folding. Folding begins co-
translationally, while the ribosome still synthesizes the polypeptide. Molecular chaperones begin to
interact with the nascent multi-domain protein as soon as it emerges from the ribosome. Co-
translational folding and chaperone interactions are recognized as crucial for efficient multi-domain
protein folding. However, these processes remain poorly defined at the molecular level, because it is
technically challenging to study them. The goal of this project is to define principles of co-translational
folding and chaperone function to better understand how complex multi-domain proteins robustly reach
their functional structures. We are using a combination of single-molecule biophysics and live-cell
experiments to accomplish this goal. With optical tweezers, we are studying the folding pathways of
nascent multi-domain proteins at the single-molecule level. Manipulation of individual molecules is
ideally suited to resolve complex folding pathways of nascent proteins, elucidate the contributions of
the ribosome and molecular chaperones to the folding process, and determine how co-translational
folding and protein synthesis are coupled and regulated to ensure robust outcomes. These detailed in
vitro studies are complemented by experiments in live cells that detect co-translational folding events
in multi-domain proteins. Protein misfolding and aggregation, misregulation of protein synthesis and
decline of chaperone function are hallmarks of many aging-related diseases. Our studies may ultimately
provide a mechanistic basis for discovering novel therapeutic strategies to treat some of these
diseases.

## Key facts

- **NIH application ID:** 11099216
- **Project number:** 3R01GM121567-08S1
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** VINCENT J. HILSER
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $149,773
- **Award type:** 3
- **Project period:** 2017-02-01 → 2026-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/11099216

## Citation

> US National Institutes of Health, RePORTER application 11099216, Folding and Chaperone Interactions of Multi-domain Proteins (3R01GM121567-08S1). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/11099216. Licensed CC0.

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