Ligand-directed acyl imidazole chemistry for labeling of membrane-bound proteins on live cells

Journal of the American Chemical Society
Sho-hei FujishimaItaru Hamachi

Abstract

Chemistry-based protein labeling in living cells is undoubtedly useful for understanding natural protein functions and for biological/pharmaceutical applications. Here, we report a novel approach for endogenous membrane-bound protein labeling for both in vitro and live cell conditions. A moderately reactive alkyloxyacyl imidazole (AI) assisted by ligand-binding affinity (ligand-directed AI (LDAI)) chemistry allowed us to selectively modify natural proteins, such as dihydrofolate reductase (DHFR) and folate receptor (FR), neither of which could be efficiently labeled using the recently developed ligand-directed tosylate approach. It was clear that LDAI selectively labeled a single Lys(K32) in DHFR, proximal to the ligand-binding pocket. We also demonstrate that the fluorescein-labeled (endogenous, by LDAI) FR works as a fluorescent biosensor on the live KB cell surface, which allowed us to carry out unprecedented in situ kinetic analysis of ligand binding to FR.

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Citations

Jun 12, 2012·Accounts of Chemical Research·Takahiro Hayashi, Itaru Hamachi
Jun 7, 2013·Medicinal Research Reviews·Ling ZhangCheng-He Zhou
Aug 16, 2016·Nature Chemical Biology·Kei YamauraItaru Hamachi
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