Screening of N-alkyl-cyanoacetamido oximes as substitutes for N-hydroxysuccinimide

Abstract

Peptide-bond formation is a pivotal process in the synthesis of peptide oligomers. Among the various coupling methodologies described, carbodiimides combine strong acylation potency and smooth reaction conditions, and they are commonly used in the presence of N-hydroxylamine additives. In recent years, acidic oxime templates, mainly ethyl 2-cyano-2-(hydroxyimino) acetate (Oxyma), have emerged as highly reactive alternatives to the classic and explosive-prone benzotriazolic additives, 1-hydroxybenzotriazole (HOBt) and 1-hydroxy-7-azabenzotriazole (HOAt). However, to achieve certain biochemical targets, less reactive species, such as N-hydroxysuccinimide (HOSu) esters, are often required to obtain stability under aqueous conditions. In the present study, we report on a new family of water-soluble N-alkyl-cyanoacetamido oximes, most of which have proven useful in the construction of active carbonates for the introduction of fluorenylmethoxycarbonyl (Fmoc) with minimal impact of dipeptide impurities. We performed a direct comparison of these new N-alkyl-cyanoacetamido oximes with HOSu in order to evaluate their capacity to retain optical purity and their coupling efficiency in the assembly of bulky residues.

Keywords: N-hydroxysuccinimides; additives; oximes; peptide syntheses; peptide-bond formations.

Figure 1

Structure of N-hydroxylamines currently available for use as additives to carbodiimides.

Figure 2

Structure of the N-alkyl-cyanoacetamido oximes proposed herein.

Scheme 1

Two-step synthetic route to cyanooxime, DmOx (9). Reagents and conditions: a) ethyl cyanoacetate (10), aq Me₂NH (40 %), 70 °C; b) Intermediate 11, NaNO₂, MeOH/H₂O, 2 h at RT.

Figure 3

Peptide models used to test the reduction of epimerization induced by N-alkyl-cyanoacetamido oximes.

Figure 4

Aib-enkephaline pentapeptide (15) used as coupling efficiency model to test N-alkyl-cyanoacetamido oximes 5–9.