REDOX-ACTIVE BIS-CYSTEINYL PEPTIDES .1. SYNTHESIS OF CYCLIC CYSTINYL PEPTIDES BY CONVENTIONAL METHODS IN SOLUTION AND ON SOLID SUPPORTS

Cyclic mono-cystinyl active-site fragments of thioredoxin and thioredo xin reductase were synthesized as N-acetyl and C-amide octapeptides by conventional methods of peptide synthesis in solution and on solid su pports. Using a side-chain protection based on acidlabile tert-butanol -derived groups and on the S-tert-butylthio unsymmetric disulfide for the thiol functions, in combination with N-alpha-Z- or N-alpha-Nps der ivatives in the chain elongation steps, the synthesis in solution was carried out in straightforward manner yielding the fully protected oct apeptides as well characterized compounds. Upon deprotection with trif luoroacetic acid and reduction of the unsymmetrical disulfides with tr i-butylphosphine, the resulting bis-cysteinyl-octapeptides were oxidiz ed in dimethylformamide with azodicarboxylic acid di-tert-butyl ester to produce the desired cyclic compounds in good overall yields. For th e synthesis on solid supports a similar acid-labile side-chain protect ion was applied in combination with the N-alpha 9-flourenylmethyoxycar bonyl derivatives in the chain elongation steps. Thereby acylations we re performed with the related amino acid N-carboxyanhydrides (UNCAs) o r by the yluronium-tetrafluoroborate/1-hydroxybenzotriazole (TBTU/HOBt ) procedure. The solid phase synthesis of the two octapeptides led to unexpected difficulties in terms of recovery of peptidic material from the resins in the final acidolytic cleavage step as well as of racemi zation at the level of the cysteine residues by the TBTU/HOBt coupling method. Racemization was efficiently suppressed by employing the rela ted pentafluorophenyl ester and this method led to crude octapeptide p roducts of a degree of purity comparable to those obtained by the synt hesis in solution. However, the recovery of the peptides from the resi n, i.e., irreversible reattachment of cleaved peptidic material via al kylation of various sidechain functions, could not be avoided even usi ng the most efficient scavengers or their cocktails. (C) 1994 John Wil ey and Sons, Inc.