Full color palette of fluorescent D-amino acids for in situ labeling of bacterial cell walls

Fluorescent D-amino acids(FDAAs) enable efficient in situ labeling of peptidoglycanin diverse bacterial species. Conducted by enzymes involved in peptidoglycanbiosynthesis, FDAA labeling allows specific probing of cell wall formation/remodeling activity, bacterial growthand cell morphology. Their broad application and high biocompatibility have made FDAAs an important and effective tool for studies of peptidoglycan synthesisand dynamics, which, in turn, has created a demand for the development of new FDAA probes. Here, we report the synthesis of new FDAAs, with emission wavelengths that span the entire visible spectrum. We also provide data to characterize their photochemical and physical properties, and we demonstrate their utility for visualizing peptidoglycan synthesisin Gram-negative and Gram-positive bacterial species. Finally, we show the permeability of FDAAs toward the outer-membrane of Gram-negative organisms, pinpointing the probes available for effective labeling in these species. This improved FDAA toolkit will enable numerous applications for the study of peptidoglycanbiosynthesis and dynamics.