Peptides, cyclic thioester synthesis

In this method, the cysteine-thioester cyclization generates a cyclic peptide 86a (see Scheme 23) with a Xaa-Cys bond whose thiol moiety is then used for tethering to the core through an S-alkylation reaction.191 The requirement for the cyclization reaction is a linear precursor 84 containing both an N-terminal Cys and a C-terminal thioester. Such a peptide precursor can be conveniently synthesized by a stepwise solid-phase synthesis on a thioester resin 81 using Boc chemistry (Scheme 22). Cleavage by HF after assembly of the peptide sequence will produce the desired precursor with an N-terminal Cys and a C-terminal thioester 84. The crude peptide is then purified by RP-HPLC and the purified unprotected peptide is then circularized in aqueous conditions buffered at pH > 7.0. 

Scheme 23 Synthesis of Cyclic Peptide Building Blocks through End-to-End Cysteine-Thioester Cycliza-tion,87n61...

Figure 14.9 Synthesis of cyclic peptides, thioesters and p-nitroanilides using the BAL linker.

The isolated TE domain from the tyrocidine (tyc) NRPS has recently been shown to catalyze the macrocyclization of unnatural substrates to generate a variety of cyclic peptides. In conjunction with standard solid-phase peptide synthesis, Walsh and coworkers demonstrated a broad substrate tolerance for peptidyl-N-acetylcysteamine thioesters by the tyrocidine TE [41,42], Cyclization of peptide analogs, where individual amino acids were replaced with ethylene glycol units, was observed with high efficiency. In addition, hydroxyacid starter units were readily cyclized by the isolated TE domain to form nonribosomal peptide-derived macrolactones. More recently, Walsh and coworkers have demonstrated effective cyclization of PEGA resin-bound peptide/polyketide hybrids by the tyrocidine TE domain [43], Utilization of a pantetheine mimic for covalent attachment of small molecules to the resin, serves as an appropriate recognition domain for the enzyme. As peptide macrocyclizations remain challenging in the absence of enzymatic assistance, this approach promises facile construction of previously unattainable structures. 

The oxidation-reduction method, developed initially by Mukaiyama et al. [133] and related to the previously described organophosphorus methods, has permitted a variety of important solid-phase applications. The mechanism of the activation is complex and involves the oxidation of the triaryl/ alkyl-phosphine to the oxide as well as reduction of the disulfide to the mercapto derivative. However, different active species, such as 81 (Fig. 11), the 2-pyridyl thioester, or even the symmetrical anhydride, have been postulated to form. For the intermediate 81, the peptide bond formation may proceed through a (cyclic transition state. The method has been used for conventional stepwise synthesis [134], acylation of the first protected amino acid to a hydroxymethyl resin, and to achieve segment condensation on a solid support in the opposite direction (N C) [135,136]. Lastly, it has been used for efficient grafting of a polyethylene glycol (molecular weight 2000) derivative to an aminomethyl resin to prepare PEG-PS resins [137]. 

The constrained conformation of cyclic peptides often results in increased exo-and e (iopeptidase resistance, enhanced binding affinity, and in certain cases, increased cell penetration compared to their linear counterparts. Numerous strategies, both in solution and solid-phase, have been reported for the synthesis of cyclic peptides [56-58] NCL, the reaction of a C-terminal peptide thioester with an... 

The synthesis of branched peptides using masked side-chain thioester derivatives of Asp and Glu which are compatible with Fmoc-SPPS is an important goal. Boll et al. synthesized cyclic and branched chain peptides using bis (2-sulfanylethyl)amido (SEA) side-chain derivatives of Asp and Glu via Fmoc SPPS [77]. The tail-to-side-chain cyclization via an in situ reduction of both acyclic and cyclic disulfides with tris(2-carboxyethyl)phosphine (TCEP) triggered the SEA intramolecular ligation. Glu derivatives cyclized more readily than the Asp analogues and without formation of side products (Scheme 9). 

Taichi M, Hemu X, Qiu YB, Tam JP (2013) A thioethylalkylamido (TEA) thioester surrogate in the synthesis of a cyclic peptide via a tandem acyl shift. Org Lett 15 2620-2623... 

Hemu X, Taichi M, Qiu YB, Liu DX, Tam JP (2013) Biomimetic synthesis of cyclic peptides using novel thioester surrogates. Biopolymers 100 492—501... 

Figure 7. General scheme of nonribosomal peptide synthesis (NRPS). Each NRPS module incorporates one amino acid into the growing peptide chain. The modules are composed of several domains Adenylation domain (red) is responsible for substrate selectivity, peptidyl carrier protein domain (orange) and condensation domain (green) work synergistically to form the peptide bond, and thioester domain (blue) which terminates the reaction, resulting in either a linear or cyclic polypeptide.

In an earlier study, we pointed out some important aspects of aminolysis of thioester in our enzyme model. First, the fastest rate for aminolysis of thioester was obtained in the presence of equimolar amounts of acid and base catalysts. Second, the reaction proceeded in aprotic nonpolar solvents such as benzene, ethyl acetate, dichloromethane, and so on [7]. Thus, the peptide syntheses by the enzyme model have been performed in benzene buffered with equimolar amounts of pivalic acid and triethylamine as acid and base catalysts, respectively. Third, the superiority of intramolecular aminolysis over an intermolecular one was clearly demonstrated, despite the large membered cyclic intermediate expected for the intramolecular reaction. The host 10 could achieve the synthesis of the tetrapep-tide derivative (11) by formal turnover of the intra-complex thiolysis and the intramolecular aminolysis, but its efficiency as an enzyme model has remained to be improved [3]. 

Thioesters can also be used for intramolecular ligation for the synthesis of end-to-end cyclic peptides (35, 36). The linear peptide precursor should contain both an N-terminal cysteine residue and a C-terminal thioester. Under previously described conditions, intramolecular thiol-exchange produces a thiolactone, which then undergoes S - N migration to yield an end-to-end cyclic peptide. 

Many other handles exist that are cleavable by orthogonal mechanisms for the synthesis of protected peptide fragments these are also summarized in Table 5. A novel strategy for the synthesis of cyclic peptides on resin is to attach the peptide to the resin by a backbone nitrogen rather that the C -carboxyl. The N-and C-termini then remain free for further functionalization such as cyclization, esterification, or thioester formation. This backbone amide linker (BAL) is listed in Table 5. Other strategies such as attaching the peptide to the resin via the side chain of trifunctional amino acids and more comprehensive listings of linkers and handles are covered in References 9,12, and 109-111. 

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