Disulfide-Driven Cyclic Peptide Synthesis of Human Endothelin-2 with a Solid-Supported Npys-Cl

Abstract

We report here the synthesis of human endothelin-2, a peptide of 21 amino acid residues with two disulfide bonds, based on the novel idea of a disulfide-driven cyclic peptide synthesis (DdCPS). This synthesis has two steps: (1) a one-pot solid-phase disulfide ligation of two different sulfur-containing peptide fragments using an Npys-Cl resin and (2) intramolecular cyclization of the disulfide peptide via amide bond formation using a thioester ligation. Human endothelin-2 was obtained in a total yield of 2.2% with two such DdCPS procedures and subsequent deprotection and HPLC purification. This strategy is the basis of a new solid-phase assisted practical synthesis of cyclic disulfide peptides.

Ready to Use Cysteine Thiol Protecting Groups in SPPS, A Practical Overview

“…This minimizes the chances of disulfide scrambling in multidisulfide peptides . The use of the Cys­(Npys) derivative facilitates efficient disulfide bond formation both in solution , and on resin mode. , While acting as an activating group, the Cys thiol of a peptide sequence is reprotected with Npys after the efficient removal of the S -protection (typically acid labileMmt) of that Cys residue, and then in the presence of a free Cys thiol the reprotected Cys­(Npys) undergoes thiol-disulfide exchange to afford the desired disulfide bond. Npys showed stability against strong acidic conditions (TFA and HF) and as a mixed disulfide-based S -protection was removed by disulfide reducing agents like BME .…”

Establishment of One-Pot Disulfide-Driven Cyclic Peptide Synthesis with a 3-Nitro-2-pyridinesulfenate

“…After the prior formation of the disulfide cross-linking between two peptide fragments by SPDSL, the intramolecular cyclization is assisted by the proximity effect of each reaction center, and more efficient and higher regioselectivity can be expected than in the intermolecular amide bond formation. 12,13) We have achieved practical syntheses of oxytocin and human endothelin-2 by DdCPS, which indicates its utility as a synthetic method to produce cyclic disulfide peptides. 12,14) However, since these syntheses involve in-solution reactions of intramolecular cyclization and deprotection of N-terminal Fmoc-group and other protecting groups, extensive HPLC purification is necessary to remove the reagents and by-products after these reactions.…”

Chemoselective Disulfide Formation by Thiol-Disulfide Interchange in SIT-Protected Cysteinyl Peptides

“…In peptide synthesis, this exchange is associated with the use of Cys-activating disulfide-based thiol-protecting groups, like 2-pyridylsulfenyl (Pyr), 3-nitro-2-pyridylsulfenyl (Npys), , and alkylsulfonyl. , The use of Cys (Npys) mediates efficient disulfide bond formation irrespective of solution , or on-resin mode. , However, the main drawback of this protecting group is its lability under basic conditions, making it incompatible with the Fmoc/ t Bu methodologythe approach most widely used in the solid phase synthesis protocol (SPPS). Moreover, commercially available Cys­(Npys) derivatives are expensive, thus limiting their use in large-scale syntheses.…”

“…6,10 The use of Cys (Npys) mediates efficient disulfide bond formation irrespective of solution 11,12 or onresin mode. 13,14 However, the main drawback of this protecting group is its lability under basic conditions, 15 making it incompatible with the Fmoc/tBu methodologythe approach most widely used in the solid phase synthesis protocol (SPPS). Moreover, commercially available Cys-(Npys) derivatives are expensive, thus limiting their use in large-scale syntheses.…”