Fluorescent tocopherols as probes of inter-vesicular transfer catalyzed by the α-tocopherol transfer protein

Novel fluorescent analogues of alpha-tocopherol have been prepared that incorporate the useful fluorophores nitrobenoxadiazyl (NBD) and anthroyloxy (AO). Both fluorescent tocopherol analogues bind specifically to recombinant human tocopherol transfer protein (hTTP). The NBD-alpha-tocopherol is particularly useful for protein-binding assays, whereas the AO-alpha-tocopherol was designed to be one of a pair of chromophores for a fluorescence resonance energy transfer (FRET) assay of intervesicular tocopherol transfer. It is now possible to follow AO-alpha-tocopherol transfer from donor lipid vesicles composed of predominantly phosphatidy1choline (PC) to acceptor lipid vesicles containing PC and a quenching lipid NBD-PE (2-dipaimitoyi-sn-glycero-3-phosphoethanolamine-N-[7-nitro-2-1,3-benzoxadiazo1-4-y1]). The presence of hTTP substantially increases the rate of AO-alpha-tolcopherol transfer over the uncatalyzed spontaneous rate.