(1)Efficient synthesis of native disulfide-rich peptides
Disulfide-rich peptides containing three or more disulfide bonds are promising therapeutic and diagnostic agents, but their preparation is often limited by the tedious and low-yielding folding process. We devised an efficient one-pot synthesis of peptides containing up to 4 disulfide bonds (snake toxin Mambalgin-1) using an azide switch strategy combined with hydrazide-based native chemical ligation. This method provides access to multi-milligram quantities of disulfide-rich peptides.
Figure 1. Synthesis of Mambalgin-1 using the azide switch strategy combined with hydrazide-based native chemical ligation.
(2) Diaminodiacid bridges to improve folding of disulfide-rich peptides
We found that a single cystine-to-diaminodiacid replacement could significantly increase the folding efficiency of disulfide-rich peptides and thus improve their production yields. The practicality of this strategy was demonstrated by the synthesis and folding of derivatives of the m-conotoxin SIIIA, the preclinical hormone hepcidin, and the trypsin inhibitor EETI-II. NMR and X-ray crystallography studies confirmed that these derivatives of disulfide-rich peptide retained the correct three-dimensional conformations. Moreover, the cystine-to-diaminodiacid replacement enabled structural tuning, thereby leading to an EETI-II derivative with higher bioactivity than the native peptide.
Reference:
-----------------------------------------------------------------------------------
1. Cui, H. K. et al. Diaminodiacid-based solid-phase synthesis of peptide disulfide bond mimics. Angewandte Chemie52, 9558-9562, doi:10.1002/anie.201302197 (2013).
2. Pan, M. et al. One-pot hydrazide-based native chemical ligation for efficient chemical synthesis and structure determination of toxin Mambalgin-1. Chem Commun (Camb), doi:10.1039/c4cc00779d (2014).
3. Guo, Y. et al. Diaminodiacid Bridges to Improve Folding and Tune the Bioactivity of Disulfide-Rich Peptides. Angewandte Chemie54, 14276-14281, doi:10.1002/anie.201500699 (2015).