The deselenization of selenocysteine selectively removes the selenol group to give alanine under anaerobic conditions or serine under aerobic conditions (oxygen saturation).

Selenocysteine, the selenium-containing analogue of cysteine, is the twenty-first proteinogenic amino acid. Since its discovery almost fifty years ago, it has been exploited in unnatural systems even more often than in natural systems. Selenocysteine chemistry has attracted the attention of many chemists in the field of chemical biology owing to its high reactivity and resulting potential for various applications such as chemical modification, chemical protein (semi)synthesis, and protein folding, to name a few. In this Minireview, we will focus on the chemistry of selenium and selenocysteine and their utility in protein chemistry.

The human body contains 25 selenoproteins, but challenges in their preparations have prevented biological characterizations thus far. Here we report the first total chemical syntheses of two human selenoproteins, selenoprotein M (SELM) and selenoprotein W (SELW).

The folding mechanism of the model protein bovine pancreatic trypsin inhibitor was revisited. By switching the solvent exposed disulfide bond with methylene thioacetal we uncovered a hidden pathway in its folding mechanism. In addition, this moiety enhanced protein stability while fully maintaining the protein structure and biological function.

The in vitro oxidative folding of proteins has been studied for over sixty years, providing critical insight into protein folding mechanisms. Hirudin, the most potent natural inhibitor of thrombin, is a 65-residue protein with three disulfide bonds, and is viewed as a folding model for a wide range of disulfide-rich proteins. Hirudin’s folding pathway is notorious for its highly heterogeneous intermediates and scrambled isomers, limiting its folding rate and yield in vitro. Aiming to overcome these limitations, we undertake systematic investigation of diselenide bridges at native and non-native positions and investigate their effect on hirudin’s folding, structure and activity. Our studies demonstrate that, regardless of the specific positions of these substitutions, the diselenide crosslinks enhanced the folding rate and yield of the corresponding hirudin analogues, while reducing the complexity and heterogeneity of the process. Moreover, crystal structure analysis confirms that the diselenide substitutions maintained the overall three-dimensional structure of the protein and left its function virtually unchanged. The choice of hirudin as a study model has implications beyond its specific folding mechanism, demonstrating the high potential of diselenide substitutions in the design, preparation and characterization of disulfide-rich proteins.

Biofilms are aggregates of microbial cells that form on surfaces and at interfaces, and are encased in an extracellular matrix. In biofilms made by the soil bacterium Bacillus subtilis, the protein TapA mediates the assembly of the functional amyloid protein TasA into extracellular fibers, and it anchors these fibers to the cell surface. We used circular dichroism and NMR spectroscopy to show that, unlike the structured TasA, TapA is disordered. In addition, TapA is composed of two weakly interacting domains: a disordered C‐terminal domain and a more structured N‐terminal domain. These two domains also exhibited different structural changes in response to changes in external conditions, such as increased temperatures and the presence of lipid vesicles. Although the two TapA domains weakly interacted in solution, their cooperative interaction with lipid vesicles prevented disruption of the vesicles. These findings therefore suggest that the two‐domain composition of TapA is important in its interaction with single or multiple partners in the extracellular matrix in biofilms.

Human SELENOF is an endoplasmic reticulum (ER) selenoprotein that contains the redox active motif CXU (C is cysteine and U is selenocysteine), resembling the redox motif of thiol-disulfide oxidoreductases (CXXC). Like other selenoproteins, the challenge in accessing SELENOF has somewhat limited its full biological characterization thus far. Here we present the one-pot chemical synthesis of the thioredoxinlike domain of SELENOF, highlighted by the use of Fmocprotected selenazolidine, native chemical ligations and deselenization reactions. The redox potential of the CXU Supporting information for this article is available on the WWW under

We describe a molecular characterization of the interaction between the cancer‐related proteins WWOX and p73. This interaction is mediated by the first of two WW domains (WW1) of WWOX and a PPXY‐motif‐containing region in p73. While phosphorylation of Tyr33 of WWOX and association with p73 are known to affect apoptotic activity, the quantitative effect of phosphorylation on this specific interaction is determined here for the first time. Using ITC and fluorescence anisotropy, we measured the binding affinity between WWOX domains and a p73 derived peptide, and showed that this interaction is regulated by Tyr phosphorylation of WW1. Chemical synthesis of the phosphorylated domains of WWOX revealed that the binding affinity of WWOX to p73 is decreased when WWOX is phosphorylated. This result suggests a fine‐tuning of binding affinity in a differential, ligand‐specific manner: the decrease in binding affinity of WWOX to p73 can free both partners to form new interactions.