Premium
Monitoring Endocytic Trafficking of Anthrax Lethal Factor by Precise and Quantitative Protein Labeling
Author(s) -
Zheng Siqi,
Zhang Gong,
Li Jie,
Chen Peng R.
Publication year - 2014
Publication title -
angewandte chemie
Language(s) - English
Resource type - Journals
eISSN - 1521-3757
pISSN - 0044-8249
DOI - 10.1002/ange.201403945
Subject(s) - bioorthogonal chemistry , endocytic cycle , anthrax toxin , cytosol , chemistry , chromosomal translocation , live cell imaging , microbiology and biotechnology , intracellular , fluorescence , cytoplasm , fusion protein , green fluorescent protein , biophysics , membrane , biochemistry , endocytosis , cell , biology , gene , combinatorial chemistry , enzyme , physics , quantum mechanics , click chemistry , recombinant dna
Coupling the genetic code expansion technique with bioorthogonal reactions enables precise control over the conjugation site as well as the choice of fluorescent probes during protein labeling. However, the advantages of this strategy over bulky and rigid fluorescent proteins (FPs) remain to be fully explored. Here we applied site‐specific bioorthogonal labeling on anthrax lethal factor (LF) to visualize its membrane translocation inside live cells. In contrast to the previously reported FP tags that significantly perturbed LF’s membrane trafficking, our precisely and quantitatively labeled LF exhibited an endocytic activity comparable to wild‐type LF. This allowed time‐lapse imaging of LF’s natural translocation process from host cell membrane to cytosol, which revealed molecular details of its virulence mechanism. Our strategy is generally applicable for monitoring intracellular protein membrane translocation that is difficult to access using conventional protein labeling methodologies.