Kunitz fold is one of the most evolutionarily ancient and conserved structural motifs, which is widely distributed among peptides of both venomous terrestrial and marine organisms. Peptides of the Kunitz/BPTI family contain about 50–70 amino acid residues cross linked by two or three disulfide bridges, adopt the conserved structural fold with two antiparallel β-sheets and one or two helical regions. Usually the peptides of this family from snakes, spiders form combinatorial libraries, the members of which have point mutations in the sequence that do not affect the spatial structure of the molecule but lead to the diversification of biological activities.
Using classical chromatography isolation, proteomic and molecular biological techniques, including NGS we discovered that tropical sea anemones of Heteractis genus produce a pool of Kunitz peptides as well. The pharmacological potential of eight peptides from combinatorial library was evaluated. All tested peptides demonstrated serine protease inhibitory activity. Five peptides showed cytoprotective activity significantly increasing the viability of neuronal Neuro-2a cells treated with 6-OHDA or β-amyloid 1-42 that could be explained by a prominent reduction in ROS production. HCRG21, the first peptide blocker of TRPV1, as well as HCRG1 and HCRG2, first Kunitz peptides from sea anemones blocking Kv1.3 channels, demonstrated anti-inflammatory effects in a mice model of carrageenan induced analgesic of paw edema. Besides, ELISA analysis showed that they effectively reduced the production of TNF-α. Our investigation shows the importance of single amino acid residues of sea anemone Kunitz-type peptides, so similar in their amino acid sequences and so different toward molecular targets. Furthermore peptides with the Kunitz fold could be seen as promising compounds for drug design as well as valuable tools for the investigation of the channels biochemistry and pharmacology.
This work was partially supported by the grants of RSF no 19-74-20088 and RFBR no 20-54-05006.