Evidence map›Paper›PMID 40306474›Full record

ArticleProtein expression and purification2025

Overcoming fluorescence loss in mEOS-based AAA+ unfoldase reporters through covalent linkage.

Isabella R Walter, Baylee A Smith, Dominic Castanzo, Matthew L Wohlever

Abstract read
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In one paragraph

Article in Protein expression and purification, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.

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0citing papers in PubMed
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1 · What the graph read from it

What it found

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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.

2 · The registry

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3 · Its place in the literature

Who cites it

0 citing papers in PubMed.

No citing paper in PubMed yet.

4 · The record

Corrections and comments

5 · Who and what money

Authors and funding

4 authors.

Isabella R WalterDepartment of Chemistry & Biochemistry, University of Toledo, Toledo, OH, 43606, USA.
Baylee A SmithDepartment of Chemistry & Biochemistry, University of Toledo, Toledo, OH, 43606, USA.
Dominic CastanzoDepartment of Molecular and Cell Biology, University of California, Berkeley, CA, 94720, USA; Institute of Quantitative Biosciences, University of California, Berkeley, CA, 94720, USA.
Matthew L WohleverDepartment of Chemistry & Biochemistry, University of Toledo, Toledo, OH, 43606, USA; Department of Cell Biology, University of Pittsburgh, Pittsburgh, PA, 15261, USA. Electronic address: wohlever@pitt.edu.

Funding

Mechanistic Investigation of Proteostasis at the Outer Mitochondrial MembraneR35GM137904 · NIGMS · UNIVERSITY OF TOLEDO · PI Matthew Lee Wohlever · 2020 to 2026
$2.6M
NIGMS NIH HHS R35 GM137904
6 · The paper itself

Abstract

Recent work has demonstrated that the soluble photoconvertable fluorescent protein mEOS can be a reporter for AAA+ (ATPases Associated with diverse cellular Activities) unfoldase activity. Given that many AAA+ proteins process membrane proteins, we sought to adapt mEOS for use with membrane protein substrates. However, direct genetic fusion of mEOS to a membrane protein completely abolished fluorescence, severely limiting the utility of mEOS for studying AAA+ proteins. To circumvent this challenge, we separately purified mEOS and multiple different AAA+ degrons, including a transmembrane domain. We then covalently linked mEOS and the degrons via Sortase. This innovative approach preserves mEOS fluorescence and photoconversion, even upon linkage to a transmembrane domain. Together, this work offers a broadly applicable platform for the study of membrane associated AAA+ proteins.

Indexed as

ATPases Associated with Diverse Cellular ActivitiesBacterial ProteinsLuminescent ProteinsMembrane ProteinsAminoacyltransferasesCysteine EndopeptidasesEscherichia coliFluorescenceRecombinant Fusion ProteinsAminoacyltransferasesATPases Associated with Diverse Cellular ActivitiesBacterial ProteinsCysteine EndopeptidasesLuminescent ProteinsMembrane ProteinsRecombinant Fusion ProteinsAAA+ proteinFluorescent proteinMembrane proteinProtein engineeringProteostasis

Identifiers

PMID40306474
PMCPMC13186246

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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.