The genetically encoded amino acid selenocysteine is predominantly found in the active sites of enzymes where it provides high chemical reactivity and specificity. The family of selenoproteins is affiliated with the management of reactive oxygen species (ROS) and provides critical health benefits associated with protecting the cell against ROS induced damage. Selenoprotein T (SelT) is a membrane-bound member of the family that resides in the endoplasmic reticulum. It was shown to contribute to cell adhesion and intracellular Ca2+ signaling through regulation of calcium channels. However, SelT has not been characterized biochemically and thus its putative enzymatic reaction leading to these functions is yet unknown. In this work, the human SelT gene has been cloned into an expression vector and the selenocysteine mutated to cysteine to facilitate its expression. SelT was expressed in E. coli at high levels and its identity was confirmed using Western Blot techniques. Subsequently it was purified using affinity chromatography. The purified SelT protein will be used in future research to determine SelT’s enzymatic reaction as well as its structure and how it is anchored in the endoplasmic reticulum’s membrane.

Membrane-bound selenoprotein T

251st National Meeting (2016)

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