Molecular determinants of heritable vitamin E deficiency

Area of Science:

• Biochemistry
• Molecular Biology
• Human Genetics

Background:

• Tocopherol transfer protein (TTP) is crucial for vitamin E homeostasis, facilitating tocopherol transport between cellular compartments.
• TTP is believed to function by enabling tocopherol secretion to the extracellular space.
• Recombinant TTP exhibits significant ability to transfer tocopherol between lipid bilayers.

Purpose of the Study:

• To biochemically characterize six missense mutations in TTP found in human AVED patients.
• To assess the impact of these mutations on TTP's affinity for RRR-alpha-tocopherol and its transfer activity between membranes.
• To investigate the correlation between in vitro TTP activity and the clinical severity of AVED.

Main Methods:

• Expression and purification of six TTP mutants (H101Q, A120T, R192H, R59W, E141K, R221W) in E. coli.
• Biochemical characterization of wild-type and mutant TTP proteins.
• Assays measuring tocopherol binding affinity and transfer kinetics between lipid membranes.

Main Results:

• Mutations R59W, E141K, and R221W, linked to severe AVED, showed impaired tocopherol binding and transfer in vitro.
• Despite severe clinical effects, these mutations had a relatively minor impact on TTP activity in vitro (2-3 fold reduction).
• Mutations associated with milder AVED forms exhibited in vitro activity similar to wild-type TTP, despite causing significant in vivo vitamin E perturbations.

Conclusions:

• In vitro tocopherol transfer activity may not accurately reflect TTP's physiological function in vivo.
• AVED syndrome might result from defects in TTP functions beyond simple tocopherol binding or transfer.
• Further research is needed to identify the specific non-transfer related activities of TTP that are affected in AVED.