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Quantitative interrogation of protein co-aggregation using multi-color fluorogenic protein aggregation sensors

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Abstract

Co-aggregation of multiple pathogenic proteins is common in neurodegenerative diseases but deconvolution of such biochemical process is challenging. Herein, we developed a dual-color fluorogenic thermal shift assay to simultaneously report on the aggregation of two different proteins and quantitatively study their thermodynamic stability during co-aggregation. Expansion of spectral coverage was first achieved by developing multi-color fluorogenic protein aggregation sensors. Orthogonal detection was enabled by conjugating sensors of minimal fluorescence crosstalk to two different proteins via sortase-tag technology. Using this assay, we quantified shifts in melting temperatures in a heterozygous model protein system, revealing that the thermodynamic stability of wild-type proteins was significantly compromised by the mutant ones but not vice versa. We also examined how small molecule ligands selectively and differentially interfere with such interplay. Finally, we demonstrated these sensors are suited to visualize how different proteins exert influence on each other upon their co-aggregation in live cells.

Graphical abstract: Quantitative interrogation of protein co-aggregation using multi-color fluorogenic protein aggregation sensors

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Supplementary files

Article information


Submitted
25 Feb 2021
Accepted
19 May 2021
First published
20 May 2021

This article is Open Access
All publication charges for this article have been paid for by the Royal Society of Chemistry

Chem. Sci., 2021, Advance Article
Article type
Edge Article

Quantitative interrogation of protein co-aggregation using multi-color fluorogenic protein aggregation sensors

Y. Bai, W. Wan, Y. Huang, W. Jin, H. Lyu, Q. Xia, X. Dong, Z. Gao and Y. Liu, Chem. Sci., 2021, Advance Article , DOI: 10.1039/D1SC01122G

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