The Allocam protecting group allows for selective deprotection and disulfide bond formation on-resin in one step.
The synthesis of disulfide-bridged peptides is a well-known challenge in peptide chemistry. When performed in solution, high dilutions have to be used in order to avoid intermolecular SS-bond formation. The on-resin formation of disulfide bonds has the advantage to favor intramolecular disulfide formation. Key on-resin methods include the treatment of peptides bearing (Ph)Acm-, Trt- or Mmt-protected cysteines with excess iodine, which may be problematic if electron-rich protecting groups are present (Postma and Albericio 2014, Eur. J. Org. Chem.
). Acidic cleavage of benzyl-type protecting groups such as Tmob, and subsequent oxidation of the resulting free thiols (e.g. using DMSO) also affords the corresponding disulfide bond. Another rather labor-intensive method for on-resin disulfide formation starts with the reductive cleavage of an StBu group, followed by oxidation with 2,2′-dithiobis(5-nitropyridine) (DTNP) to form RS–SNP. A single Mmt group is then cleaved under acidic conditions, and the resulting free thiol forms the desired SS-bond by microwave-assisted displacement of the SNP group (Galanis et al. 2009, Pept. Sci.
Key methods for on-resin disulfide bond formation.
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The Allocam-protecting group, which is a variant of the Acm protecting group, allows for a palladium-mediated single-step approach using mild reaction conditions and readily available reagents. In a recently published synthesis of the C-carboxy analog of oxytocin (Kondasinghe et al.
2017, Org. Biomol. Chem.
), complete conversion was achieved on Aminomethyl ChemMatrix® resin using 1.5 equivalents of Pd(OAc) 2 as catalyst, together with 3% NMM as well as 5% AcOH (v/v). No reduced peptide intermediates or S-allylated products were observed, and no residual palladium was detected in the final product. The reaction proceeds using the protected resin-bound peptide without loss of any acid-labile side chain protecting groups. By using Fmoc-L-Cys(Allocam)-OH, solution-phase manipulations of the peptide can be avoided, and the number of HPLC purification steps can be minimized.
- Direct palladium-mediated on-resin disulfide formation from Allocam protected peptides; T. D. Kondasinghe, H. Y. Saraha, S. B. Odeesho and J. L. Stockdill; Org Biomol Chem 2017; 15: 2914-2918. doi:10.1039/c7ob00536a
- Disulfide Formation Strategies in Peptide Synthesis; T. M. Postma and F. Albericio; European Journal of Organic Chemistry 2014; 2014: 3519-3530. doi:10.1002/ejoc.201402149
- Enhanced microwave-assisted method for on-bead disulfide bond formation: synthesis of alpha-conotoxin MII; A. S. Galanis, F. Albericio and M. Grotli; Peptide Science 2009; 92: 23-34. doi:10.1002/bip.21116
- Allylic protection of thiols and cysteine: I: The allyloxycarbonylaminomethyl group; A. Malanda Kimbonguila, A. Merzouk, F. Guibé and A. Loffet; Tetrahedron 1999; 55: 6931-6944. doi:10.1016/s0040-4020(99)00322-1
- Allylic protecting groups and their use in a complex environment part II: Allylic protecting groups and their removal through catalytic palladium π-allyl methodology; F. Guibé; Tetrahedron 1998; 54: 2967-3042. doi:10.1016/s0040-4020(97)10383-0
- Use of Alloc-amino acids in solid-phase peptide synthesis. Tandem deprotection-coupling reactions usingneutral conditions; N. Thieriet, J. Alsina, E. Giralt, F. Guibé and F. Albericio; Tetrahedron Letters 1997; 38: 7275-7278. doi:10.1016/s0040-4039(97)01690-0
- The allyloxycarbonylaminomethyl group: a new allytic protection for the thiol group of cysteine; A. M. Kimbonguila, A. Merzouk, F. Guibé and A. Loffet; Tetrahedron Letters 1994; 35: 9035-9038. doi:10.1016/0040-4039(94)88420-x
- S-2,4,6-trimethoxybenzyl (Tmob): a novel cysteine protecting group for the N.alpha.-(9-fluorenylmethoxycarbonyl) (Fmoc) strategy of peptide synthesis; M. C. Munson, C. Garcia-Echeverria, F. Albericio and G. Barany; The Journal of Organic Chemistry 1992; 57: 3013-3018. doi:10.1021/jo00037a013