Targeting marine jellyfish toxins: development of metalloproteinase inhibitors through a specific method for protease...

The venom of hazardous jellyfish contains harmful substances to human health, such as metalloproteinases, which are challenging to purify. Therefore, identifying substrate sequences using crude venom is crucial for the development of peptide inhibitors. This study utilized a designed peptide library and an abundance-based hydrolysis product analysis approach to identify the substrate sequences of toxin metalloproteinases from Nemopilema nomurai nematocyst venom (NnNV) and further modified these sequences into peptide inhibitors. Specifically, a peptide library comprising 41 potential substrate sequences was constructed and incubated with NnNV. 12 substrate sequences and 14 cleavage sites were identified from the hydrolysis products. In the design of peptide inhibitors, thiol (-SH) and phosphate groups (-PO₃H₂) were used as zinc-binding groups, and sixty derived peptides were obtained. 15 peptide inhibitors were selected for their efficacy in inhibiting toxin metalloproteinase activity, with CPRGQPIIQDV demonstrating the most potent effect. CPRGQPIIQDV mainly presented β-sheet and random coil structures, and significantly reduced the cytotoxicity and pro-inflammatory effects of NnNV on RAW264.7 cells. This study established a method for identifying metalloproteinase substrate sequences using crude jellyfish venom and provided an initial design and screening of peptide inhibitors, offering new insights into the management of jellyfish stings.