Phosphatidylserine (PS), an anionic lipid with the amino acid L-serine in the hydrophilic head group is present predominantly in the inner leaflet of mammalian cell membranes and could reach concentrations of up to 30% of the total lipid content.  Disruption of the asymmetry of phosphatidylserine in which it translocates from the inner to the outer leaflet and gets surface exposed occurs with the onset of diseases like cancer and Alzheimer's and when cells become apoptotic.  We have developed novel fluorescent protein probes employing the C2 domain of lactadherin in which two small fluorescent molecules such as NBD (nitorbenzodioxazole) are attached to adjacent cysteines (Lact-C2-Cys-NBD probe) or lysines (Lact-C2-Lys-NBD probe) to target PS.  The unqiue properties of the Lact-C2-NBD probes are: (1) the NBD molecules are proximal and exciton coupled with a red fluorescence (680 nm maximum) when the probes are not PS bound, (2) the NBD molecules become distal when the probes bind PS and  exhibit a green fluorescence (540 nm maximum) characteristic of individual NBD molecules without exciton coupling, (3) they are membrane permeable as they can bind to PS in the outer and inner leaflets of DOPC/DOPS vesicle and cell membranes of live prostate cells without the addition of any membrane permeabilizing or PS translocating reagents, and (4) they have excellent phtostability.  Significant aspects of this research will be presented.

Exciton coupled membrane permeable fluorescent probes for phosphatidylserine

243rd National Meeting (2012)

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