Open AccessDesign, Synthesis, and Application of the Trimethoprim‐Based Chemical Tag for Live‐Cell Imaging
Author(s) -
Jing Chaoran,
Cornish Virginia W.
Publication year - 2013
Publication title -
current protocols in chemical biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.503
H-Index - 14
ISSN - 2160-4762
DOI - 10.1002/9780470559277.ch130019
Subject(s) - trimethoprim , chemistry , computational biology , biology , computer science , microbiology and biotechnology , antibiotics
Over the past decade, chemical tags have been developed to complement the use of fluorescent proteins in live‐cell imaging. Chemical tags retain the specificity of protein labeling achieved with fluorescent proteins through genetic encoding, but provide smaller, more robust tags and modular use of organic fluorophores with high photon output and tailored functionalities. The trimethoprim‐based chemical tag (TMP‐tag) was initially developed based on the high affinity interaction between E. coli dihydrofolate reductase and the antibiotic trimethoprim and was subsequently rendered covalent and fluorogenic via proximity‐induced protein labeling reactions. To date, the TMP‐tag is one of the few chemical tags that enable intracellular protein labeling and high‐resolution live‐cell imaging. Here we describe the general design, chemical synthesis, and application of TMP‐tag for live‐cell imaging. Alternate protocols for synthesizing and using the covalent and the fluorogenic TMP‐tags are also included. Curr. Protoc. Chem. Biol . 5:131‐155 © 2013 by John Wiley & Sons, Inc.