Selenoprotein K form an intermolecular diselenide bond with unusually high redox potential

Abstract

Selenoprotein K (SelK) is a membrane protein involved in antioxidant defense, calcium regulation and the ER-associated protein degradation pathway. We found that SelK exhibits a peroxidase activity with a rate that is low but within the range of other peroxidases. Notably, SelK reduced hydrophobic substrates, such as phospholipid hydroperoxides, which damage membranes. Thus, SelK might be involved in membrane repair or related pathways. SelK was also found to contain a diselenide bond — the first intramolecular bond of that kind reported for a selenoprotein. The redox potential of SelK was −257 mV, significantly higher than that of diselenide bonds in small molecules or proteins. Consequently, SelK can be reduced by thioredoxin reductase. These finding are essential for understanding SelK activity and function.

Selenoprotein K and Protein Palmitoylation

“…As discussed earlier, purified SelK can dimerize via an intermolecular diselenide bond that exhibits strong reducing potential (22). Purified SelK exhibits some level of peroxidase activity and is able to reduce hydrophobic substrates such as phospholipid hydroperoxides, which is consistent with antioxidant activity displayed by human SelK overexpressed in cells (23).…”

“…The cytosolic region of SelK is rich in both prolines and glycines, with a calpain-2 cleavage site present in the cytosolic region between amino-acid residues Arg81 and Gly82 (17). Purified human SelK was found to dimerize through the formation of intermolecular diselenide bonds (22), but it remains unclear whether this occurs in the ER membrane of mammalian cells. Other than the Sec residue, the most distinctive feature of SelK is a predicted Src homology 3 (SH3) binding domain.…”

“…SelK can bind to ERAD components, including Derlin-1 and the p97 ATPase, and is involved in transport of glycosylated misfolded proteins and regulation of ER stress in some cell lines (9,38). Overexpression of SelK in cardiomyocytes has been implicated in potential antioxidant responses (23), and purified human SelK was shown to reduce phospholipid hydroperoxides (22). This may suggest a role for SelK in ER membrane repair, although it remains to be determined whether this occurs in vivo.…”

Preparation of Selenocysteine-Containing Forms of Human SELENOK and SELENOS

“…It also interacts with additional p97 adaptors, substrate recruiters, and partners [9]. In contrast to SELENOS, SELENOK’s specific role in the ERAD pathway is not yet known [1, 10]. In addition to its involvement in the ERAD, SELENOK, however, was shown to bind and stabilize the palmitoyl transferase DHHC6 [11].…”

“…Furthermore, the two proteins are intrinsically disordered and as such they are prone to proteolytic cleavage in host organisms [15]. All these challenges were addressed by expressing both SELENOK and SELENOS in the robust host E. coli using a fused protein partner to increase solubility and prevent integration into E. coli membranes [10, 16–18]. For both the proteins the incorporation of Sec is essential to maintain their functional integrity [10, 11, 17, 18].…”