A systematic approach to the solid-phase synthesis of linear and cyclic pseudopeptide libraries containing psi[CH2NH] amide bond surrogates.

Abstract

A systematic approach has been adopted for the synthesis and characterization of a series of linear and cyclic pseudopeptide mixtures containing the psi[CH2NH] amide replacement. The parent structures were based on biologically relevant compounds including an enkephalin analog, H-Tyr-D-Ala-Gly-Phe-Leu-OH, and an Arg-Gly-Asp peptide sequence. The linear mixtures containing 4 and 64 pseudopeptide components with 1, 2 or 3 amide bond surrogates were synthesized using Boc-SPPS. The amount of desired linear pseudopeptides in the mixtures ranged from 67 to 90% as determined by integration of HPLC peak areas. Comparative studies indicated: (i) racemization is not a problem in the synthesis of pseudopeptide mixtures containing the psi[CH2NH] surrogate; and (ii) protection of the psi[CH2NH] surrogate with a benzyloxycarbonyl group during the synthesis is beneficial. Cyclic mixtures containing 4 and 256 cyclic pseudopeptide components with a single amide bond surrogate were synthesized using a resin-bound cyclization approach featuring side-chain attachment of Boc-Asp-OFm to the solid support. Cyclization kinetic studies revealed that the newly developed HATU coupling reagent provided a fast cyclization rate for a pseudopeptide mixture and that the position of the reduced peptide bond within a peptide mixture had only a small effect on the cyclization rates of the mixture. Pseudopeptide libraries permit the more efficient bioassay of complex structures and can also be used to reveal more rapidly trends in physicochemical variables. For example, we observed that the expected increase in hydrophilicity with psi[CH2NH] substitutions during RP-HPLC analysis did not continue with several such replacements.