Folding stabilities of ribosome-bound nascent polypeptides probed by mass spectrometry

The folding of most proteins occurs during the course of their translation while their tRNA-bound C termini are embedded in the ribosome. How the close proximity of nascent proteins to the ribosome influences their folding thermodynamics remains poorly understood. Here, we have developed a mass spec...

Full description

Saved in:
Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 2023-08, Vol.120 (33), p.e2303167120-e2303167120
Main Authors: Tan, Ruiyue, Hoare, Margaret, Welle, Kevin A, Swovick, Kyle, Hryhorenko, Jennifer R, Ghaemmaghami, Sina
Format: Article
Language:English
Subjects:
Biological Sciences
Charge distribution
Chemotaxis
Dihydrofolate reductase
DNA polymerase
DNA-directed DNA polymerase
Electrostatic properties
Folding
Mass Spectrometry
Mass spectroscopy
Methionine
Oxidation
Peptides - chemistry
Polypeptides
Protein Biosynthesis
Protein Folding
Proteins
Proteins - metabolism
Proteomics
Reductases
Ribosomes - metabolism
Scientific imaging
Stability analysis
Thermodynamics
tRNA
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:The folding of most proteins occurs during the course of their translation while their tRNA-bound C termini are embedded in the ribosome. How the close proximity of nascent proteins to the ribosome influences their folding thermodynamics remains poorly understood. Here, we have developed a mass spectrometry-based approach for determining the stabilities of nascent polypeptide chains using methionine oxidation as a folding probe. This approach enables quantitative measurement subglobal folding stabilities of ribosome nascent chains within complex protein mixtures and extracts. To validate the methodology, we analyzed the folding thermodynamics of three model proteins (dihydrofolate reductase, chemotaxis protein Y, and DNA polymerase IV) in soluble and ribosome-bound states. The data indicate that the ribosome can significantly alter the stability of nascent polypeptides. Ribosome-induced stability modulations were highly variable among different folding domains and were dependent on localized charge distributions within nascent polypeptides. The results implicated electrostatic interactions between the ribosome surface and nascent polypeptides as the cause of ribosome-induced stability modulations. The study establishes a robust proteomic methodology for analyzing localized stabilities within ribosome-bound nascent polypeptides and sheds light on how the ribosome influences the thermodynamics of protein folding.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.2303167120