RADA

RADA is a fluorescent D-amino acid (FDAA) with high photostability and thermostability, which emits yellow-to-orange fluorescence. RADA shows low outer membrane permeability in wild-type Gram-negative Escherichia coli, but it targets penicillin-binding proteins and L,D-transpeptidases, mimics the interaction between acyl acceptors and enzyme intermediates, and integrates into peptidoglycan during biosynthesis. As a peptidoglycan labeling reagent, RADA metabolically integrates into the nascent peptidoglycan of live bacterial cells, labels the peptidoglycan at the poles and lateral walls of mycobacteria, and enables visualization of peptidoglycan synthesis and remodeling processes. RADA serves as a non-specific stain for fixed cells, is non-toxic to bacterial cells, and its red-shifted excitation/emission spectra reduce phototoxicity. RADA also supports virtual pulse-chase labeling experiments and stochastic optical reconstruction microscopy for sub-diffraction-limited imaging of bacterial cell walls^[1].