A growth-based platform for detecting domain-peptide interactions in the cytoplasm of mammalian cells
Development of a method for detecting protein-protein interactions (PPIs) in living cells is important for therapeutic drug screening against various diseases including infectious diseases. We have recently developed a method named SOS localization-based interaction screening (SOLIS), in which we de...
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Published in | Scientific reports Vol. 12; no. 1; pp. 18028 - 10 |
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Main Authors | , , |
Format | Journal Article |
Language | English |
Published |
England
Nature Publishing Group
27.10.2022
Nature Portfolio |
Subjects | |
Online Access | Get full text |
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Summary: | Development of a method for detecting protein-protein interactions (PPIs) in living cells is important for therapeutic drug screening against various diseases including infectious diseases. We have recently developed a method named SOS localization-based interaction screening (SOLIS), in which we designed membrane-anchored and SOS-fused chimeric proteins, whose PPI-dependent association triggers membrane localization of the SOS-fused chimeric protein, activates the Ras/MAPK pathway, and induces cell growth. While SOLIS was able to detect relatively strong PPIs, further sensitivity was required for detecting intracellular endogenous PPIs typically having a micromolar order of dissociation constant (K
). Here we develop high-sensitive SOLIS (H-SOLIS) that could universally detect PPIs with lower affinities. In order to improve the sensitivity, H-SOLIS introduces a heterodimeric helper interaction, in which addition of a small-molecule helper ligand could accommodate association of the two chimeric proteins and regulate the sensitivity. Four types of domain-peptide interactions having known K
values are employed to examine the versatility and detection limit of H-SOLIS. Consequently, the heterodimer-inducible helper ligand dramatically enhances detection sensitivity, lowering the detection limit to a ten-micromolar order of K
. Thus, H-SOLIS could be a platform to detect disease-related domain-peptide interactions for drug discovery screening. |
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ISSN: | 2045-2322 2045-2322 |
DOI: | 10.1038/s41598-022-22770-4 |