A chemical tool for blue light-inducible proximity photo-crosslinking in live cells.
Pratyush Kumar MishraMyeong-Gyun KangHakbong LeeSeungjoon KimSubin ChoiNirmali SharmaCheol-Min ParkJaewon KoChangwook LeeJeong Kon SeoHyun-Woo RheePublished in: Chemical science (2021)
We developed a proximity photo-crosslinking method ( Spotlight ) with a 4-azido- N -ethyl-1,8-naphthalimide (AzNP) moiety that can be converted to reactive aryl nitrene species using ambient blue light-emitting diode light. Using an AzNP-conjugated HaloTag ligand (VL1), blue light-induced photo-crosslinked products of various HaloTag-conjugated proteins of interest were detected in subcellular spaces in live cells. Chemical or heat stress-induced dynamic changes in the proteome were also detected, and photo-crosslinking in the mouse brain tissue was enabled. Using Spotlight , we further identified the host interactome of SARS-CoV-2 nucleocapsid (N) protein, which is essential for viral genome assembly. Mass analysis of the VL1-crosslinked product of N -HaloTag in HEK293T cells showed that RNA-binding proteins in stress granules were exclusively enriched in the cross-linked samples. These results tell that our method can reveal the interactome of protein of interest within a short distance in live cells.
Keyphrases
- induced apoptosis
- sars cov
- stress induced
- light emitting
- cell cycle arrest
- photodynamic therapy
- air pollution
- signaling pathway
- oxidative stress
- gene expression
- genome wide
- cell death
- respiratory syndrome coronavirus
- small molecule
- dna methylation
- electron transfer
- coronavirus disease
- heat stress
- amino acid
- pi k akt
- protein protein