A Chemical Tool for Blue Light-Inducible Proximity Photo-crosslinking in Live Cells

ABSTRACT: 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. 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.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Permanent Cell Line Cell

SUBMITTER: Hyun-Woo Rhee  

LAB HEAD: Hyun-Woo Rhee

PROVIDER: PXD027149 | Pride | 2022-04-04

REPOSITORIES: Pride