Plant Antimicrobial Peptides Snakin-1 and Snakin-2: Chemical Synthesis and Insights into the Disulfide Connectivity

Antimicrobial peptides and proteins represent an important class of plant defensive compounds against pathogens and provide a rich source of lead compounds in the field of drug discovery. We describe the effective preparation of the cysteine‐rich snakin‐1 and ‐2 antimicrobial peptides by using a combination of solid‐phase synthesis and native chemical ligation. A subsequent cysteine/cystine mediated oxidative folding to form the six internal disulfide bonds concurrently gave the folded proteins in 40–50 % yield. By comparative evaluation of mass spectrometry, HPLC, biological data and trypsin digest mapping of folded synthetic snakin‐2 compared to natural snakin‐2, we demonstrated that synthetic snakin‐2 possesses full antifungal activity and displayed similar chromatographic behaviour to natural snakin‐2. Trypsin digest analysis allowed tentative assignment of three of the purported six disulfide bonds. Snakin potatoes: An efficient chemical synthesis of the disulfide‐rich antimicrobial potato proteins snakin‐1 and snakin‐2 is reported (see figure). The synthetic proteins were compared to the naturally occurring snakins by HPLC and mass spectrometric analyses, and the antifungal activity was found to be equally potent. Trypsin cleavage of synthetic and natural snakin‐2 allowed partial assignment of the disulfide pattern.