Total chemical synthesis of seleno-hirudins and their oxidative folding studies

The in vitro oxidative folding has been extensively studied in the last four decades especially with small disulfide-rich proteins. One conclusion that these studies provide is that many disulfide-rich proteins follow two different folding model mechanisms; those seen for bovine pancreatic trypsin inhibitor (BPTI) and for hirudin, or a combination of the two opposite folding models. Selenocysteine (Sec) incorporation has been successfully used in protein folding studies including that of BPTI. Sec`s low redox potential, and pK_a, as well as its increased nucleophilicity and electrophilicity can enhance thiol-disulfide-like exchange reactions that are essential for protein folding. Based on these results we wish to study the effect of Sec substitution on the folding of hirudin. Wild-type hirudin and its seleno-analogs containing one or two substitutions at different positions that could form native diselenide bonds have been prepared using solid phase peptide synthesis (SPPS) and native chemical ligation (NCL). Here we present preliminary data that shows the distinct difference in the oxidative folding of Hirudin and its seleno analogs.