Enzyme‐Catalyzed Peptide‐Bond Formation: Elastase‐ and δ‐Chymotrypsin‐Assisted Synthesis of Oligopeptides

Abstract

A series of penta-and hexapeptides of the structure Ac-L,-. . .-LxI-LyI-. . . -Lym-NH, with phenylalanine or tyrosine in L,, and glycine or alanine in the other positions were synthesized using elastase and d-chymotrypsin to catalyse the formation of the peptide bonds adjacent to the aromatic residue. Such oligopeptides are useful substrates for studying the secondary specificity of chymotrypsin-like proteases.Introduction'). -Growing interest in protease-catalyzed synthesis of peptides is documented by a large number of model studies showing the mode of application of various proteases and by several reports on the successful use of this approach in the synthesis of biologically active peptides. Comprehensive reviews of these investigations have been published [3] [4]. Further examples of preparative peptide synthesis for which the enzymatic approach presents clear advantages over the conventional methods will help recognising cases in which the enzymatic method may open valuable alternative synthetic routes.In our laboratory, enzyme-catalyzed formation of peptide bonds has become a key step in the synthesis of numerous peptide substrates we need for our investigation of the specificity of serine proteases. The systematical variation of the peptide structure required by these studies is achieved by synthesizing various small peptides as building blocks and then assembling them in different combinations. According to a general strategy, the small fragments are prepared stepwise by conventional methods and then assembled using proteases as catalysts, thus assuring racemization-free fragment condensation. In relation to our study of the interactions of the extended active site of a-chymotrypsin, we were let to design a series of peptide substrates containing an internal aromatic residue to specify the scissible peptide bond and several adjacent alanine or glycine residues. The 6-chymotrypsin-assisted synthesis of a number of such peptides has been described in a previous communication [2]. In the present paper we report on the synthesis of further members of this series, the pentapeptides Ac-Ala-Ala-Ala-Tyr-Gly-NH, (lx-ly) and Ac-Ala-Ala-Ala-Tyr-Ala-NH, (lx-2y) and the hexapeptides Ac-Ala-Ala-Ala-Tyr-Ala-

Direct Transacylation of 2,2,2-Trihaloethyl Esters with Amines and Alcohols Using Phosphorus(III) Reagents for Reductive Fragmentation and in Situ Activation

“…N - tert -Butoxycarbonyl- l -alanyl- l -alanine Ethyl Ester (9). Amide 9 was obtained in 70% yield (39.1 mg) from 6 and l -alanine ethyl ester hydrochloride as a white solid (flash chromatography 50% EtOAc in hexanes). Data for 9 : R f 0.38 (60% EtOAc in hexanes); [α] 22 D −48.1 ( c = 1.0, EtOH); IR (thin film) 3332, 3261, 3075, 2948, 1752, 1688, 1656, 1540, 1521, 1269, 1167 cm -1 ; 1 H NMR (CDCl 3 , 300 MHz) δ 6.73 (br s, 1H), 5.12 (br s, 1H), 4.55 (quin, 1H, J = 7.4 Hz), 4.23−4.16 (m, 3H), 1.45 (s, 9H), 1.40 (d, 3H, J = 7.2 Hz), 1.37 (d, 3H, J = 7.2 Hz), 1.28, (t, 3H, J = 7.2 Hz); 13 C NMR (CDCl 3 , 75 MHz) δ 172.7 (C), 172.2 (C), 155.4 (C), 79.5 (C).…”