Bulky Aspartate Protecting Groups to Avoid Aspartimide Formation

Bulky Aspartate Protecting Groups to Avoid Aspartimide Formation

Published on 09/03/2021

Frustrated by aspartimide formation during peptide synthesis? Iris Biotech presents aspartate derivatives with bulky side chain protecting groups that minimize the formation of aspartimide-related side-products.

Advances in the field of peptide synthesis regarding coupling agents, the availability of pre-coupled building blocks, and optimized resins enable the suppression of racemization and lead to increased product yields.

Although peptide chemistry is constantly developing, the process of aspartimide formation during peptide synthesis remains a serious challenge limiting the accessibility of certain peptide sequences. Aspartimides are formed upon ring-closure between the nitrogen of the alpha-carboxyl amide bond and the ß-carboxyl sidechain and release of the carboxyl protecting group. As this side reaction is promoted by strong bases such as piperidine which is commonly used for Fmoc removal, this problem is especially pronounced during Fmoc SPPS.

The formed aspartimides undergo rapid epimerization followed by ring opening either by hydrolysis or by virtue of base, thus leading to the formation of up to nine potential by-products. Consequently, aspartimide formation leads to low products yields in addition to time- and cost-intensive purification. As some of those by-products are even co-eluting on HPLC due to identical retention times compared to the desired product, they may be virtually impossible to remove rendering certain peptide sequences inaccessible.

 

 

Aspartimide formation and possible side products.

Aspartimide formation can be decreased by increasing the steric bulk of the aspartic acid ester moiety. Iris Biotech offers bulky beta-trialkyl-methyl ester protected aspartic acid building blocks, which allow to minimize aspartimide formation during Fmoc SPPS of Asp-containing peptides.

 

Bulky Aspartate derivatives offered by Iris Biotech helping to avoid aspartimide formation.

Analogues of the tert-butyl group in which the methyl group is substituted for longer alkyl chains help to shield the aspartyl beta-carboxyl group and thereby reduce the formation of aspartimide-derived by-products. Our bulky side-chain protecting groups provide considerably more protection against the formation of aspartimide-related by-products than the commonly used OtBu group.

 

References:

  • Preventing aspartimide formation in Fmoc SPPS of Asp-Gly containing peptides - practical aspects of new trialkylcarbinol based protecting groups; R. Behrendt, S. Huber, P. White; J. Pept. Sci. 2016; 22(2): 92-97. https://doi.org/10.1002/psc.2844.
  • New t-butyl based aspartate protecting groups preventing aspartimide formation in Fmoc SPPS; R. Behrendt, S. Huber, R. Marti, P. White; J. Pept. Sci. 2015; 21(8): 680-687. https://doi.org/10.1002/psc.2790.
  • 2-phenyl isopropyl esters as carboxyl terminus protecting groups in the fast synthesis of peptide fragments; C. Yue, J. Thierry, P. Potier; Tetrahedron Lett. 1993; 34: 323-326. https://doi.org/10.1016/S0040-4039(00)60578-6.
  • The aspartimide problem in Fmoc-based SPPS. Part I; M. Mergler, F. Dick, B. Sax, P. Weiler, T. Vorherr; J. Pept. Sci. 2003; 9(1): 36-46. https://doi.org/10.1002/psc.430.
  • A new protecting group for aspartic acid that minimizes piperidine-catalyzed aspartimide formation in Fmoc solid phase peptide synthesis; A. Karlström, A. Undén; Tetrahedron Lett. 1996; 37(24): 4243-4246. https://doi.org/10.1016/0040-4039(96)00807-6.
  • The aspartimide problem in Fmoc‐based SPPS. Part II; M. Mergler, F. Dick, B. Sax, C. Stähelin, T. Vorherr; J. Pept. Sci. 2003; 9(8): 518-526. https://doi.org/10.1002/psc.473.
  • Prevention of aspartimide formation during peptide synthesis using cyanosulfurylides as carboxylic acid-protecting groups; K. Neumann, J. Farnung, S. Baldauf, J. W. Bode; Nat. Commun. 2020; 11: 982. https://doi.org/10.1038/s41467-020-14755-6.
  • Patent EP 2 886 531 B1
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