STUDIES OF THE TRANSMEMBRANE SELENOPROTEIN K AND ITS ROLE IN OXIDATIVE DEFENSE

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Selenium is an essential element in all branches of life, where its strong nucleophilicity, redox activity, metal binding capacities, and low pKa are utilized to catalyze redox and electron transfer reactions. In human health, selenium compounds and selenoproteins contribute to chemopreventive, anti-inflammatory, and antiviral defense. This project focuses on elucidating the unknown structure and function of the selenium-containing membrane protein, selenoprotein K (SelK). SelK is a small (94 residues) protein with a single-pass transmembrane helix predicted at its N terminus, and a selenocysteine at its C terminus. It has been shown to reduce internal levels of reactive oxygen species and to protect cells against oxidative stress. Oxidative stress, caused by externally and internally generated reactive oxygen species (ROS), is an unavoidable corollary of aerobic life and is tightly connected to longevity. ROS tend to concentrate in the lipid bilayer where oxygen and other free radicals are more soluble. Not surprisingly, ROS are prone to modify both lipids and membrane proteins, with lipid peroxidation being the most common ROS-induced damage. Such modified lipids change the physico-chemical properties, e.g. fluidity and curvature stress of the lipid bilayer, and thus destabilize biomembranes. Due to the specialized cellular roles of selenoproteins, SelK's participation in anti-oxidant defense in vivo and SelK's membrane localization, we hypothesize that SelK is involved in protecting biomembranes against reactive carbonyl species generated by lipid oxidation. To facilitate its structural and biophysical characterization we are developing bacterial overexpression and purification strategies for both the native selenium and sulfur-substituted SelK. The oligomerization state of SelK and sample conditions for structural characterization will be evaluated. To identify the role of SelK in oxidative stress and signaling, we will use functional assays to test for its potential peroxidase activity. Our long-term goal is to delineate the biochemical role of SelK, the unique contribution of selenium to its reactivity, and SelK function in its native membrane environment.