Synthesis and characterization of bichromophoric 1-deoxyceramides as FRET probes.

The suitability as FRET probes of the bichromophoric 1-deoxydihydroceramides RBM5-160 and RBM5 161, containing NBD spisulosine as sphingoid base and a terminal MCC or NR tagged palmitic acid, respectively, has been evaluated. Moreover, the ceramide synthase (CerS) catalyzed metabolic incorporation of ω azido palmitic acid into NBD spisulosine to render the corresponding ω azido 1 deoxyceramide RBM5-159 has been studied in several cell lines. In addition, the SPAAC reaction between RBM5 159 and the BCN derived NR or MCC fluorophores has been optimized to render the target bichromophoric 1 deoxydihydroceramides. These results pave the way for the development of FRET based assays related to sphingolipid metabolism.