A thiol probe for measuring unfolded protein load and proteostasis in cells.
Nat Commun
; 8(1): 474, 2017 09 07.
Article
in En
| MEDLINE
| ID: mdl-28883394
ABSTRACT
When proteostasis becomes unbalanced, unfolded proteins can accumulate and aggregate. Here we report that the dye, tetraphenylethene maleimide (TPE-MI) can be used to measure cellular unfolded protein load. TPE-MI fluorescence is activated upon labelling free cysteine thiols, normally buried in the core of globular proteins that are exposed upon unfolding. Crucially TPE-MI does not become fluorescent when conjugated to soluble glutathione. We find that TPE-MI fluorescence is enhanced upon reaction with cellular proteomes under conditions promoting accumulation of unfolded proteins. TPE-MI reactivity can be used to track which proteins expose more cysteine residues under stress through proteomic analysis. We show that TPE-MI can report imbalances in proteostasis in induced pluripotent stem cell models of Huntington disease, as well as cells transfected with mutant Huntington exon 1 before the formation of visible aggregates. TPE-MI also detects protein damage following dihydroartemisinin treatment of the malaria parasites Plasmodium falciparum. TPE-MI therefore holds promise as a tool to probe proteostasis mechanisms in disease.Proteostasis is maintained through a number of molecular mechanisms, some of which function to protect the folded state of proteins. Here the authors demonstrate the use of TPE-MI in a fluorigenic dye assay for the quantitation of unfolded proteins that can be used to assess proteostasis on a cellular or proteome scale.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Sulfhydryl Compounds
/
Molecular Probes
/
Cells
/
Protein Folding
/
Proteostasis
Limits:
Animals
/
Humans
Language:
En
Journal:
Nat Commun
Journal subject:
BIOLOGIA
/
CIENCIA
Year:
2017
Document type:
Article
Affiliation country:
Australia