Abstract:
The a-tocopherol transfer protein (a-TTP) is responsible for the retention of the atocopherol
form of vitamin E in living organisms. The detailed ligand transfer
mechanism by a-TTP is still yet to be fully elucidated. To date, studies show that a-TTP
transfers a-tocopherol from late endosomes in liver cells to the plasma membrane where
it is repackaged into very low density lipoprotein (VLDL) and released into the
circulation.
Late endosomes have been shown to contain a lipid known as lysobisphosphatidic
acid (LBP A) that is unique to this cellular compartment. LBPA plays a role in
intracellular trafficking and controlling membrane curvature. Taking these observations
into account plus the fact that certain proteins are recruited to membranes based on
membrane curvature, the specific aim of this project was to examine the effect of LBP A
on a-TTP binding to lipid membranes. To achieve this objective, dual polarization
interferometry (DPI) and a vesicle binding assay were employed. Whilst DPI allows
protein binding affinity to be measured on a flat lipid surface, the vesicle binding assay
determines protein binding affinity to lipid vesicles mimicking curved membranes.
DPI analysis revealed that the amount of a-TTP bound to lipid membranes is
higher when LBPA is present. Using the vesicle binding assay, a similar result was seen
where a greater amount of protein is bound to large unilamellar vesicles (LUV s)
containing LBP A. However, the effect of LBP A was attenuated when small unilamellar
vesicles (SUVs) were replaced with LUVs. The outcome of this project suggests that aTTP
binding to membranes is influenced by membrane curvature, which in turn is
induced by the presence of LBP A.
