title: Electronic and Steric Optimization of Fluorogenic Probes for Biomolecular Imaging.
abstract: Fluorogenic probes are invaluable tools for spatiotemporal investigations within live cells. In common fluorogenic probes, the intrinsic fluorescence of a small-molecule fluorophore is masked by esterification until entry into a cell, where endogenous esterases catalyze the hydrolysis of the masking groups, generating fluorescence. The susceptibility of masking groups to spontaneous hydrolysis is a major limitation of these probes. Previous attempts to address this problem have incorporated auto-immolative linkers at the cost of atom economy and synthetic adversity. Here, we report on a linker-free strategy that employs adventitious electronic and steric interactions in easy-to-synthesize probes. We find that X···C═O n→π* interactions and acyl group size are optimized in 2',7'-dichlorofluorescein diisobutyrate. This probe is relatively stable to spontaneous hydrolysis but is a highly reactive substrate for esterases both in vitro and in cellulo, yielding a bright, photostable fluorophore with utility in biomolecular imaging.
authors.affiliation: Department of Chemistry, ‡Graduate Program in Biophysics, and §Department of Biochemistry, University of Wisconsin-Madison , Madison, Wisconsin 53706, United States.
authors.forename: Wen
authors.initials: W
authors.lastname: Chyan
authors.affiliation: Department of Chemistry, ‡Graduate Program in Biophysics, and §Department of Biochemistry, University of Wisconsin-Madison , Madison, Wisconsin 53706, United States.
authors.forename: Henry R
authors.initials: HR
authors.lastname: Kilgore
authors.affiliation: Department of Chemistry, ‡Graduate Program in Biophysics, and §Department of Biochemistry, University of Wisconsin-Madison , Madison, Wisconsin 53706, United States.
authors.forename: Brian
authors.initials: B
authors.lastname: Gold
authors.affiliation: Department of Chemistry, ‡Graduate Program in Biophysics, and §Department of Biochemistry, University of Wisconsin-Madison , Madison, Wisconsin 53706, United States.
authors.forename: Ronald T
authors.identifier: 0000-0001-7164-1719
authors.initials: RT
authors.lastname: Raines
mesh_terms: D000818:Animals; D002384:Catalysis; D018929:Cell Culture Techniques; D003470:Culture Media; D005456:Fluorescent Dyes / Q000737:chemistry*; D013050:Spectrometry, Fluorescence / Q000379:methods*
chemical_list: D003470:Culture Media; D005456:Fluorescent Dyes