A NEW SHK-LIKE PEPTIDE, NNK-1, FROM THE GIANT NOMURA’S JELLYFISH DISPLAYS HKV1.3 CHANNEL BLOCKING

Abstract

Based on its genomic information, we have discovered a new human voltage-gated potassium channel (hKv1.3) blocker, NnK-1 (CKDHHTYGVY10CKDWKSSGEC20KKNPKG MRHF30 CRKTCGFC38), from the jellyfish Nemopilema nomurai. The gene sequence for NnK-1 contains 5,408 base pairs with five introns and six exons. The coding sequence of its precursor is 894 nucleotides and 297 amino acids long, containing five presumptive cystein-rich, ShK-like peptides. Among these peptides, we chemically synthesized the fifth, NnK-1, which contain three pairs of disulfide bonds (C1-C6, C2-C4, and C3-5), to investigate their bioactivity. For the electrophysiological assay, we devised an internal ribosome entry site (IRES) vector to separately express hKv1.3 and enhanced green fluorescence protein (EGFP), which we then transfected into HEK 293 cells. Our assay demonstrated that NnK-1 is an effective hKv1.3 blocker. Multiple alignment of cnidarian Shk-like peptides, which have Kv1.3 blocking activity, revealed that four residues (3Asp, 25Lys, 33Lys, and 34Thr in NnK-1) along with six cysteine residues are conserved. We therefore hypothesize that these four residues are crucial for the toxins to bind to the voltage-gated potassium channel.