title: A rapid and fluorogenic TMP-AcBOPDIPY probe for covalent labeling of proteins in live cells.
abstract: Protein labeling is enormously useful for characterizing protein function in cells and organisms. Chemical tagging methods have emerged as a new generation protein labeling strategy in live cells. Here we have developed a novel and versatile TMP-AcBOPDIPY probe for selective and turn-on labeling of proteins in live cells. A small monomeric tag, E. coli dihydrofolate reductase (eDHFR), was rationally designed to introduce a cysteine in the vicinity of the ligand binding site. Trimethoprim (TMP) that specifically binds to eDHFR was linked to the BOPDIPY fluorophore containing a mildly thiol-reactive acrylamide group. TMP-AcBOPDIPY rapidly labeled engineered eDHFR tags via a reaction termed affinity conjugation (a half-life of ca. 2 min), which is one of the top fast chemical probes for protein labeling. The probe displays 2-fold fluorescence enhancement upon labeling of proteins. We showed that the probe specifically labeled intracellular proteins in live cells without and with washing out the dye. We demonstrated its utility in visualizing intracellular processes by fluorescence-lifetime imaging microscopy (FLIM) measurements.
authors.affiliation: Chemical Genomics Centre of the Max Planck Society , Otto-Hahn-Str. 15, 44227 Dortmund, Germany.
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
authors.forename: Fu
authors.initials: F
authors.lastname: Li
authors.forename: Xi
authors.initials: X
authors.lastname: Chen
authors.forename: Jian
authors.initials: J
authors.lastname: Hou
authors.forename: Long
authors.initials: L
authors.lastname: Yi
authors.forename: Yao-Wen
authors.initials: YW
authors.lastname: Wu
mesh_terms: D002470:Cell Survival; D004926:Escherichia coli / Q000201:enzymology; D005456:Fluorescent Dyes / Q000737:chemistry*; D006367:HeLa Cells; D006575:Heterocyclic Compounds, 3-Ring / Q000737:chemistry*; D006801:Humans; D011162:Porphobilinogen / Q000031:analogs & derivatives* / Q000737:chemistry; D013194:Staining and Labeling; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry*; D013997:Time Factors; D014295:Trimethoprim / Q000031:analogs & derivatives* / Q000737:chemistry*
chemical_list: D005456:Fluorescent Dyes; D006575:Heterocyclic Compounds, 3-Ring; C000593088:TMP-AcBOPDIPY probe; C507544:dipyrromethene; D011162:Porphobilinogen; D014295:Trimethoprim; D013762:Tetrahydrofolate Dehydrogenase