Grafting bioactive peptide sequences onto small cysteine-rich scaffolds is a promising strategy for enhancing their stability and value as novel peptide-based therapeutics. However， correctly folded disulfide-rich peptides can be challenging to produce by either recombinant or synthetic means. The single disulfide-directed β-hairpin (SDH) fold， first observed in contryphan-Vc1， provides a potential alternative to complex disulfide-rich scaffolds. We have undertaken recombinant production of full-length contryphan-Vc1 (rCon-Vc1[Z1Q]) and a truncated analogue (rCon-Vc1[Z1Q])， analyzed the backbone dynamics of rCon-Vc1[Z1Q]， and probed the conformational and proteolytic stability of these peptides to evaluate the potential of contryphan-Vc1 as a molecular scaffold. Backbone N relaxation measurements for rCon-Vc1[Z1Q] indicate that the N-terminal domain of the peptide is ordered up to Thr19， whereas the remainder of the C-terminal region is highly flexible. The solution structure of truncated rCon-Vc1[Z1Q] was similar to that of the full-length peptide， indicating that the flexible C-terminus does not have any effect on the structured domain of the peptide. Contryphan-Vc1 exhibited excellent proteolytic stability against trypsin and chymotrypsin but was susceptible to pepsin digestion. We have investigated whether contryphan-Vc1 can accept a bioactive epitope while maintaining the structure of the peptide by introducing peptide sequences based on the DINNN motif of inducible nitric oxide synthase. We show that sCon-Vc1[NNN] binds to the iNOS-binding protein SPSB2 with an affinity of 1.3 μM while maintaining the SDH fold. This study serves as a starting point in utilizing the SDH fold as a peptide scaffold.