Diselenides as proxies of disulfides in cystine-rich peptides
The strongly reducing redox potential and isosteric character of selenocysteine compared to cysteine allows to install in correct position a diselenide bridge when a pair of natively-crossbridged cysteine residues are replaced synthetically by a pair of selenocysteines. This regioselective formation of the diselenide bond represents a powerful strategy for directing oxidative folding of peptides containing multiple disulfide bonds into the native diselenide/disulfide framework by reducing the number of possible disulfide isomers and thus enhancing folding efficiency. This is well documented by the successful synthesis of conotoxins, most notably of -conotoxins with two disulfide bonds, but also of 3-disulfide-bridged — and –conotoxins which generally show a low propensity to form in satisfactory yields by air oxidation of the multiple cysteine-precursors the native disulfide isomer.
Boc, tert-butyloxycarbonyl; BPTI, bovine pancreatic trypsin inhibitor; Bzl, benzyl; DTT, dithiothreitol; DMSO, dimethylsulfoxide; DTNP, 2,2´-dithiobis(5-nitropyridine); Fmoc, fluorenylmethoxycarbonyl; Grx, glutaredoxin; GSH, glutathione; GSSG, oxidized glutathione; Meb, p-methylbenzyl; Mob, p-methoxybenzyl; StBu, tert-butylsulfanyl; Sec or U, selenocysteine; trx, thioredoxin; tBu, tert-butyl; TFA, trifluoroacetic acid; Trt, trityl.
Early studies of protein chemistry led Anfinsen to propose the principle that correct oxidative folding of proteins is a thermodynamically coupled process uniquely governed by the amino acid sequence (1). The acquired experience, however, clearly revealed that yields of in vitro folding, particularly in absence of the natural folding catalysts such as chaperones and protein-disulfide isomerases, strongly depend on experimental conditions (temperature, salts, redox buffers, etc.). Correspondingly, over the last decades the related methodology has been continuously optimized such that recovery of correctly folded proteins is generally achieved in satisfactory to high yields (2). Moreov ...