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Systematic identification of structure-specific protein–protein interactions


Abstract

Protein–protein interactions (PPIs) mediate numerous essential functions and regulatory events in living organisms. The physical interactome of a protein can be abnormally altered in response to external and internal cues, thus modulating cell physiology and contributing to human disease. In particular, neurodegenerative diseases due to the accumulation of aberrantly folded and aggregated proteins may lead to alterations in protein interactomes. Identifying changes in the interactomes of normal and disease states of proteins could help to understand molecular disease mechanisms, but current interactomics methods are limited in the ability to pinpoint structure-specific PPIs and their interaction interfaces on a proteome-wide scale. Here, we adapted limited proteolysis–mass spectrometry (LiP–MS) to systematically identify putative structure-specific PPIs by probing protein structural alterations within cellular extracts upon treatment with specific structural states of a given protein. We demonstrate the feasibility of our method to detect well-characterized PPIs, including antibody–target protein interactions and interactions with membrane proteins, and show that it pinpoints PPI interfaces. We then applied the LiP–MS approach to study the structure-specific interactors of the Parkinson’s disease hallmark protein alpha-synuclein (aSyn). We identified several previously known interactors of both aSyn monomer and amyloid fibrils and provide a resource of novel putative structure-specific interactors for further studies. This approach is applicable to identify structure-specific interactomes of any protein, including posttranslationally modified and unmodified, or metabolite-bound and unbound structural states of proteins.

Abstract

Protein–protein interactions (PPIs) mediate numerous essential functions and regulatory events in living organisms. The physical interactome of a protein can be abnormally altered in response to external and internal cues, thus modulating cell physiology and contributing to human disease. In particular, neurodegenerative diseases due to the accumulation of aberrantly folded and aggregated proteins may lead to alterations in protein interactomes. Identifying changes in the interactomes of normal and disease states of proteins could help to understand molecular disease mechanisms, but current interactomics methods are limited in the ability to pinpoint structure-specific PPIs and their interaction interfaces on a proteome-wide scale. Here, we adapted limited proteolysis–mass spectrometry (LiP–MS) to systematically identify putative structure-specific PPIs by probing protein structural alterations within cellular extracts upon treatment with specific structural states of a given protein. We demonstrate the feasibility of our method to detect well-characterized PPIs, including antibody–target protein interactions and interactions with membrane proteins, and show that it pinpoints PPI interfaces. We then applied the LiP–MS approach to study the structure-specific interactors of the Parkinson’s disease hallmark protein alpha-synuclein (aSyn). We identified several previously known interactors of both aSyn monomer and amyloid fibrils and provide a resource of novel putative structure-specific interactors for further studies. This approach is applicable to identify structure-specific interactomes of any protein, including posttranslationally modified and unmodified, or metabolite-bound and unbound structural states of proteins.

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Additional indexing

Item Type:Working Paper
Communities & Collections:04 Faculty of Medicine > Institute of Medical Genetics
08 Research Priority Programs > Adaptive Brain Circuits in Development and Learning (AdaBD)
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Uncontrolled Keywords:Limited proteolysis, Mass spectrometry, Structural proteomics, Protein–protein interactions, Structure-specific interactions
Language:English
Date:3 February 2023
Deposited On:08 Feb 2023 12:08
Last Modified:09 May 2024 03:24
Series Name:ArXiv.org
Number of Pages:38
ISSN:2331-8422
OA Status:Green
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1101/2023.02.01.522707
Project Information:
  • : FunderUniversity of Zurich
  • : Grant IDURPP AdaBD
  • : Project TitleUniversity Research Priority Program AdaBD (Adaptive Brain Circuits in Development and Learning),
  • : Project Websitehttps://www.adabd.uzh.ch/
  • Content: Published Version
  • Language: English
  • Licence: Creative Commons: Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0)